Polypeptides and polynucleotides, and uses thereof as a drug target for producing drugs and biologics

ABSTRACT

This invention relates to a novel target for production of immune and non-immune based therapeutics and for disease diagnosis. More particularly, the invention provides therapeutic antibodies against VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 antigens, which are predicted co-stimulatory family members and which are differentially expressed in cancers including, lung cancer, ovarian cancer, and colon cancer, and diagnostic and therapeutic usages. The use of these antibodies for modulating B7 costimulation and related therapies such as the treatment of autoimmunity are also provided. This invention further relates to the discovery of extracellular domains of VSIG1 and its variants, FXYD3 and its variants, ILDR1 and its variants, LOC253012 and its variants, AI216611 and its variants, and C1ORF32 and its variants which are suitable targets for immunotherapy, cancer therapy, and drug development.

FIELD OF THE INVENTION

This invention relates to the discovery of certain proteins that aredifferentially expressed in specific tissues and their use astherapeutic and diagnostic targets. More specifically the inventionrelates to a protein VSIG1 and its variants, FXYD3 and its variants,ILDR1 and its variants, LOC253012 and its variants, AI216611 and itsvariants, and C1ORF32 and its variants, which are differentiallyexpressed by some cancers, and therefore are suitable targets forimmunotherapy, cancer therapy, and drug development. This inventionfurther relates to the discovery of extracellular domains of VSIG1 andits variants, FXYD3 and its variants, ILDR1 and its variants, LOC253012and its variants, AI216611 and its variants, and C1ORF32 and itsvariants which are suitable targets for immunotherapy, cancer therapy,and drug development

Additionally, because some of the proteins of this invention, based ontheir B7-like structure, are believed to play a role in immunecostimulation, the invention further relates to the use of theseproteins, or drugs which modulate these proteins (agonistic andantagonistic), as immune modulators and for immune therapy, especiallyfor treating cancer and immune related disorders such as cancers andautoimmune disorders. Also, the invention more specifically relates totherapeutic and diagnostic antibodies and therapies and diagnosticmethods using same antibodies and antibody fragments that specificallybind to proteins of invention or a soluble or secreted portion thereof,especially the ectodomain.

BACKGROUND OF THE INVENTION

Tumor antigens are ideally positioned as biomarkers and drug targets,and they play a critical role in the development of novel strategies foractive and passive immunotherapy agents, to be used as stand-alonetherapies or in conjunction with conventional therapies for cancer.Tumor antigens can be classified as either tumor-specific antigens(TSAs) where the antigens are expressed only in tumor cells and not innormal tissues, or tumor-associated antigens (TAAs) where the antigensare overexpressed in tumor cells but nonetheless also present at lowlevels in normal tissues.

TAAs and TSAs are validated as targets for passive (antibody) therapy aswell as active immunotherapy using strategies to break immune toleranceand stimulate the immune system. The antigenic epitopes that aretargeted by these therapeutic approaches are present at the cellsurface, overexpressed in tumor cells compared to non-tumor cells, andare targeted by antibodies that block functional activity, inhibit cellproliferation, or induce cell death.

There are growing number of tumor-associated antigens against whichmonoclonal antibodies have been tested or are in use as treatment forcancer. The identification and molecular characterization of novel tumorantigens expressed by human malignancies is an active field in tumorimmunology. Several approaches have been used to identifytumor-associated antigens as target candidates for immunotherapy,including high throughput bioinformatic approaches, based on genomicsand proteomics. The identification of novel TAAs or TSAs expands thespectrum of tumor antigen targets available for immune recognition andprovides new target molecules for the development of therapeutic agentsfor passive immunotherapy, including monoclonal antibodies, whetherunmodified or armed. Such novel antigens may also point the way to moreeffective therapeutic vaccines for active or adoptive immunotherapy.

Cancer vaccination involves the administration of tumor antigens and isused to break immune tolerance and induce an active T-cell response tothe tumor. Vaccine therapy includes the use of naked DNA, peptides,recombinant protein, and whole cell therapy, where the patient's owntumor cells are used as the source of the vaccine. With theidentification of specific tumor antigens, vaccinations are more oftencarried out by dendritic cell therapy, whereby dendritic cells areloaded with the relevant protein or peptide, or transfected with vectorDNA or RNA.

The major applications of anti-TAA antibodies for treatment of cancerare therapy with naked antibody, therapy with a drug-conjugatedantibody, and fusion therapy with cellular immunity. Ever since theirdiscovery, antibodies were envisioned as “magic bullets” that woulddeliver toxic agents, such as drugs, toxins, enzymes and radioisotopes,specifically to the diseased site and leaving the non-target normaltissues unaffected. Indeed, antibodies, and in particular antibodyfragments, can function as carriers of cytotoxic substances such asradioisotopes, drugs and toxins. Immunotherapy with suchimmunoconjugates is more effective than with the naked antibody.

In contrast to the overwhelming success of naked (such as Rituxan andCampath) and conjugated antibodies (such as Bexxar and Zevalin) intreating hematological malignancies, only modest success has beenachieved in the immunotherapy of solid tumors. One of the majorlimitations in successful application of immunotherapy to solid tumorsis the large molecular size of the intact immunoglobulin that results inprolonged serum half-life but in poor tumor penetration and uptake.Indeed, only a very small amount of administered antibody (as low as0.01%) reaches the tumor. In addition to their size, antibodiesencounter other impediments before reaching their target antigensexpressed on the cell surface of solid tumors. Some of the barriersinclude poor blood flow in large tumors, permeability of vascularendothelium, elevated interstitial fluid pressure of tumor stroma, andheterogenous antigen expression.

With the advent of antibody engineering, small molecular weight antibodyfragments exhibiting improved tumor penetration have been generated.Such antibody fragments are often conjugated to specific cytotoxicmolecules and are designed to selectively deliver them to cancer cells.Still, solid tumors remain a formidable challenge for therapy, even withimmunoconjugated antibody fragments.

The new wave of optimization strategies involves the use of biologicalmodifiers to modulate the impediments posed by solid tumors. Thus, incombination to antibodies or their conjugated antibody fragments,various agents are being used to improve the tumor blood flow, enhancevascular permeability, lower tumor interstitial fluid pressure bymodulating stromal cells and extracellular matrix components, upregulateexpression of target antigens and improve penetration and retention ofthe therapeutic agent.

Immunotherapy with antibodies represents an exciting opportunity forcombining with standard modalities, such as chemotherapy, as well ascombinations with diverse biological agents to obtain a synergisticactivity. Indeed, unconjugated mAbs are more effective when used incombination with other therapeutic agents, including other antibodies.

Another component of the immune system response to immunotherapy is thecellular response, specifically—the T cell response and activation ofcytotoxic T cells (CTLs). The efficiency of the immune system inmediating tumor regression depends on the induction of antigen-specificT-cell responses through physiologic immune surveillance, priming byvaccination, or following adoptive transfer of T-cells. Although avariety of tumor-associated antigens have been identified and manyimmunotherapeutic strategies have been tested, objective clinicalresponses are rare. The reasons for this include the inability ofcurrent immunotherapy approaches to generate efficient T-cell responses,the presence of regulatory cells that inhibit T-cell responses, andother escape mechanisms that tumors develop, such as inactivation ofcytolytic T-cells through expression of negative costimulatorymolecules. Effective immunotherapy for cancer will require the use ofappropriate tumor-specific antigens; the optimization of the interactionbetween the antigenic peptide, the APC and the T cell; and thesimultaneous blockade of negative regulatory mechanisms that impedeimmunotherapeutic effects.

T-cell activation plays a central role in driving both protective andpathogenic immune responses, and it requires the completion of acarefully orchestrated series of specific steps that can be preempted ordisrupted by any number of critical events. Naïve T cells must receivetwo independent signals from antigen-presenting cells (APC) in order tobecome productively activated. The first, Signal 1, is antigen-specificand occurs when T cell antigen receptors encounter the appropriateantigen-MHC complex on the APC. A second, antigen-independent signal(Signal 2) is delivered through a T cell costimulatory molecule thatengages its APC-expressed ligand. In the absence of a costimulatorysignal, T-cell activation is impaired or aborted, which may lead to astate of antigen-specific unresponsiveness (known as T-cell anergy), ormay result in T-cell apoptotic death.

Costimulatory signals can be either stimulatory (positive costimulation)or inhibitory (negative costimulation or coinhibition). Positivecostimulation is required for optimal activation of naïve T cells, whilenegative costimulation is required for the acquisition of immunologictolerance to self, as well as the termination of effector T cellfunctions. Costimulatory signals, particularly positive costimulatorysignals, also play a role in the modulation of B cell activity. Forexample, B cell activation and the survival of germinal center B cellsrequire T cell-derived signals in addition to stimulation by antigen.

Both positive and negative costimulatory signals play critical roles inthe regulation of cell-mediated immune responses, and molecules thatmediate these signals have proven to be effective targets forimmunomodulation. Based on this knowledge, several therapeuticapproaches that involve targeting of costimulatory molecules have beendeveloped, and were shown to be useful for prevention and treatment ofcancer and autoimmune diseases, as well as rejection of allogenictransplantation, each by turning on, or preventing the turning off, ofimmune responses in subjects with these pathological conditions.

Costimulatory molecule pairs usually consist of ligands expressed onAPCs and their cognate receptors expressed on T cells. The wellcharacterized B7/CD28 and CD40/CD40L costimulatory molecules arecritical in primary T-cell activation. In recent years, severaladditional costimulatory molecules have been identified, that belong tothe B7/CD28 or the TNF/TNF-R gene families. The effects of costimulatoryTNFR family members can often be functionally, temporally, or spatiallysegregated from those of CD28 family members and from each other. Thesequential and transient regulation of T cell activation/survivalsignals by different costimulators may function to allow longevity ofthe response while maintaining tight control of T cell survival.

The B7 family consists of structurally related, cell-surface proteinligands, which bind to receptors on lymphocytes that regulate immuneresponses. Interaction of B7-family members with their respectivecostimulatory receptor, usually a member of the CD28-related family,augments immune responses, while interaction with coinhibitoryreceptors, such as CTLA4, attenuates immune responses. Members of the B7family share 20-40% amino-acid identity and are structurally related,with the extracellular domain containing tandem domains related tovariable and constant immunoglobulin domains.

There are currently seven known members of the family: B7.1 (CD80), B7.2(CD86), B7-H1 (PD-L1), B7-H2 (ICOS-L), B7-DC (PD-L2), B7-H3, and B7-H4,each with unique, yet often overlapping functions. Clearly, each B7molecule has developed its own indispensable niche in the immune system.As specific niches of B7 family members continue to be dissected, theirdiagnostic and therapeutic potential becomes ever more apparent. Many ofthe B7 superfamily members were initially characterized as T cellcostimulatory molecules. However, more recently it has become clear theycan also coinhibit T cell responses. Thus, B7 family members may haveopposing effects on an immune response.

Central to the normal function of the immune system is its ability todistinguish between self and non-self, since failure to do so couldprovoke the onset of autoimmune disease. Most autoimmune disorders areknown to involve autoreactive T cells and/or autoantibodies. Thus,agents that are capable of inhibiting or eliminating autoreactivelymphocytes have a promising therapeutic potential. Furthermore, the useof agents that exhibit such immunosuppressive activity should also bebeneficial in order to inhibit normal immune responses to alloantigensin patients receiving a transplant. Thus, novel agents that are capableof modulating costimulatory signals, without compromising the immunesystem's ability to defend against pathogens, are highly advantageousfor treatment and prevention of such pathological conditions.

The importance of the B7 family members in regulating immune responsesto self and allo-antigens was demonstrated by the development ofimmunodeficiency and autoimmune diseases in mice with mutations inB7-family genes. Accordingly, manipulation of the signals delivered byB7 ligands has shown potential in the treatment of autoimmunity,inflammatory diseases, and transplant rejection. This approach relies,at least partially, on the eventual deletion of auto- or allo-reactive Tcells, presumably because in the absence of costimulation (which inducescell survival genes) T cells become highly susceptible to induction ofapoptosis.

Harnessing the immune system to treat chronic diseases is a major goalof immunotherapy. Active and passive immunotherapies are provingthemselves as effective therapeutic strategies. Passive immunotherapy,using monoclonal antibodies or receptor Fc-fusion proteins, has come ofage and has shown great clinical success. A growing number of suchtherapeutic agents have been approved or are in clinical trials toprevent allograft rejection or to treat autoimmune diseases and cancer.Active immunotherapy (i.e. vaccines) has been effective against agentsthat normally cause acute self-limiting infectious diseases followed byimmunity and has been at the forefront of efforts to prevent theinfectious diseases that plague humankind. However, active immunotherapyhas been much less effective against cancer or chronic infectiousdiseases primarily because these have developed strategies to escapenormal immune responses. Among these are negative costimulators of theB7 family, such as B7-H1 and B7-H4, which are highly expressed incertain tumors, and afford local protection from immune cells-mediatedattack.

The efficiency of the immune system in mediating tumor regressiondepends on the induction of antigen-specific T-cell responses throughphysiologic immune surveillance, priming by vaccination, or followingadoptive transfer of T-cells. Although a variety of tumor-associatedantigens have been identified and many immunotherapeutic strategies havebeen tested, objective clinical responses are rare. The reasons for thisinclude the inability of current immunotherapy approaches to generateefficient T-cell responses, the presence of regulatory cells thatinhibit T-cell responses, and other escape mechanisms that tumorsdevelop, such as inactivation of cytolytic T-cells through expression ofnegative costimulatory molecules. Effective immunotherapy for cancerwill require the use of appropriate tumor-specific antigens; theoptimization of the interaction between the antigenic peptide, the APCand the T cell; and the simultaneous blockade of negative regulatorymechanisms that impede immunotherapeutic effects.

Costimulators of the B7 family play a critical role in activation andinhibition of antitumor immune responses. Novel agents targeting thesemolecules could find significant use in the modulation of immuneresponses and the improvement of cancer immunotherapy. Such agents couldbe administered in conjunction with tumor-specific antigens, as anadjuvant that serves to enhance the immune response to the antigen inthe patient. In addition, such agents could be of use in other types ofcancer immunotherapy, such as adoptive immunotherapy, in whichtumor-specific T cell populations are expanded and directed to attackand kill tumor cells. Agents capable of augmenting such anti-tumorresponse have great therapeutic potential and may be of value in theattempt to overcome the obstacles to tumor immunotherapy.

Passive tumor immunotherapy uses the exquisite specificity and lyticcapability of the immune system to target tumor specific antigens andtreat malignant disease with a minimum of damage to normal tissue.Several approaches have been used to identify tumor-associated antigensas target candidates for immunotherapy. The identification of noveltumor specific antigens expands the spectrum of tumor antigen targetsavailable for immune recognition and provides new target molecules forthe development of therapeutic agents for passive immunotherapy,including monoclonal antibodies, whether unmodified or armed. Such novelantigens may also point the way to more effective therapeutic vaccinesfor active or adoptive immunotherapy.

Clinical development of costimulation blockade came to fruition with theapproval of CTLA4Ig (abatacept) for rheumatoid arthritis. This solublefusion protein, which acts as competitive inhibitor of the B7/CD28costimulatory pathway, is also in clinical trials for other immunediseases such as psoriasis and multiple sclerosis, and for transplantrejection. Promising results have also been obtained in a phase IIclinical trial in kidney transplantation with belatacept, are-engineered CTLA4Ig with enhanced binding affinity to its ligands,B7.1 and B7.2 (CD80 and CD86, respectively). Two fully human anti-CTLA4monoclonal antibodies, Ipilimumab and tremelimumab, abrogate theCTLA4/B7 inhibitory interaction, and are in clinical phase III formetastatic melanoma and other cancers, as well as HIV infection.Galiximab is a primatized monoclonal antibody targeting CD80, in PhaseII for rheumatoid arthritis, psoriasis and Non-Hodgkin's lymphoma.

It is important to point out that strategies that use single agents toblock costimulation have often proved to be insufficient. Given thediversity of the different costimulation molecules, future strategiesmay involve the simultaneous blockade of several selected pathways orcombination therapy with conventional drugs, such as immunosuppressantsfor immune-related disorders or cytotoxic drugs for cancer.

Despite recent progress in the understanding of cancer biology andcancer treatment, as well as better understanding of the moleculesinvolved in immune responses, the success rate for cancer therapy andfor the treatment of autoimmune diseases remains low. Therefore, thereis an unmet need for new therapies which can successfully treat bothcancer and autoimmune disorders.

BRIEF SUMMARY OF THE INVENTION

It is an object of the invention to provide novel therapeutic anddiagnostic compositions containing at least one of the VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, or FXYD3 proteins or one of the novelsplice variants disclosed herein as well as to provide these novel VSIG1splice variants; specifically ILDR1 splice variants; LOC253012 splicevariants; AI216611 splice variants, C1ORF32 splice variants; and FXYD3splice variants, and nucleic acid sequences encoding for same orfragments thereof especially the ectodomain or secreted forms of VSIG1,ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 proteins and/or splicevariants.

It is another object of the invention to use said proteins, splicevariants and nucleic acid sequences as novel targets for development ofdrugs which specifically bind to the VSIG1, ILDR1, LOC253012, AI216611,C1ORF32, FXYD3 proteins and/or splice variants, and/or drugs whichagonize or antagonize the binding of other moieties to the VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, FXYD3 proteins and/or splice variants.

It is still another object of the invention to provide drugs whichmodulate (agonize or antagonize) at least one VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 or FXYD3 related biological activity. Such drugsinclude by way of example antibodies, small molecules, peptides,ribozymes, antisense molecules, siRNA's and the like. These moleculesmay directly bind or modulate an activity elicited by the VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, FXYD3 proteins or VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, FXYD3 DNA or portions or variants thereof or mayindirectly modulate a VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3associated activity or binding of molecules to VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, FXYD3 and portions and variants thereof such as bymodulating the binding of VSIG1, ILDR1, LOC253012, AI216611, C1ORF32,FXYD3 to its counterreceptor or endogenous ligand.

In more specific embodiments, the present invention provides novelsplice variants of a known protein V-set and immunoglobulin domaincontaining 1 (SEQ ID NO:11) (RefSeq accession identifier NP_(—)872413,synonyms: RP5-889N15.1, 1700062D20Rik, GPA34, MGC44287, dJ889N15.1) or apolynucleotide encoding same, which can be used as diagnostic markersand/or therapeutic agents which agonize or antagonize the binding ofother moieties to the VSIG1 proteins and/or which modulate (agonize orantagonize) at least one VSIG1 related biological activity.

According to one more specific embodiment, the novel splice variant isan isolated polynucleotide comprising a nucleic acid having a nucleicacid sequence as set forth in any one of AI581519_T10 (SEQ ID NO:9),AI581519_T11 (SEQ ID NO:10), or a sequence homologous thereto. Accordingto another embodiment, the isolated polynucleotide is at least 95%homologous to any one of AI581519_T10 (SEQ ID NO:9), AI581519_T11 (SEQID NO:10).

According to yet another more specific embodiment, the novel splicevariant is an isolated protein or polypeptide having an amino acidsequence as set forth in any one of AI581519_P9 (SEQ ID NO:15),AI581519_P10 (SEQ ID NO:16), or a sequence homologous thereto. Accordingto another embodiment, the isolated polypeptide is at least 95%homologous to any one of AI581519_P9 (SEQ ID NO:15), AI581519_P10 (SEQID NO:16).

It is another specific object of the invention to provide molecules andisolated polypeptides comprising the soluble ectodomain (ECD) of theVSIG1 proteins and fragments thereof as well as nucleic acid sequencesencoding said soluble ectodomain, as well as fragments thereof andconjugates and the use thereof as therapeutics including their use inimmunotherapy (promoting or inhibiting immune costimulation).

In more specific embodiments the present invention provides discreteportions of the VSIG1 proteins including different portions of theextracellular domain corresponding to residues 23-234 of the VSIG1protein sequence contained in the sequence of AI581519_P3 (SEQ IDNO:11), corresponding to amino acid sequence depicted in SEQ ID NO:138,or residues 23-270 of the of the VSIG1 protein sequence contained in thesequence of AI581519_P4 (SEQ ID NO:12), corresponding to amino acidsequence depicted in SEQ ID NO:139, or residues 23-296 of the VSIG1protein sequence contained in the sequence of AI581519_P5 (SEQ IDNO:13), corresponding to amino acid sequence depicted in SEQ ID NO:140,or residues 23-193 of the VSIG1 protein sequence contained in thesequence of AI581519_P7 (SEQ ID NO:14) corresponding to amino acidsequence depicted in SEQ ID NO:141, or residues 23-203 of the VSIG1protein sequence contained in the of AI581519_P9 (SEQ ID NO:15)corresponding to amino acid sequence depicted in SEQ ID NO:142, orresidues 23-231 of the VSIG1 protein sequence contained in the sequenceof AI581519_P10 (SEQ ID NO:16), corresponding to amino acid sequencedepicted in SEQ ID NO:143, or residues 26-293 of the VSIG1 proteinsequence contained in the sequence of AI581519_P5 (SEQ ID NO:13),corresponding to amino acid sequence depicted in SEQ ID NO:302, orvariants thereof possessing at least 80% sequence identity, morepreferably at least 90% sequence identity therewith and even morepreferably at least 95, 96, 97, 98 or 99% sequence identity therewith.

According to other more specific embodiments, the present inventionprovides novel splice variants of a known protein immunoglobulin-likedomain containing receptor 1 (SEQ ID NO:21) (RefSeq accession identifierNP_(—)787120, also known as ILDR1alpha, ILDR1beta, ILDR1), or apolynucleotide encoding same, which can be used as diagnostic markersand/or therapeutic agents which agonize or antagonize the binding ofother moieties to the ILDR1 proteins and/or which modulate (agonize orantagonize) at least one ILDR1 related biological activity.

In one specific embodiment, the novel splice variant is an isolatedpolynucleotide comprising a nucleic acid having a nucleic acid sequenceas set forth in AA424839_(—)1_T7 (SEQ ID NO:20), or a sequencehomologous thereto. According to another embodiment, the isolatedpolynucleotide is at least 95, 96, 97, 98 or 99% homologous toAA424839_(—)1_T7 (SEQ ID NO:20).

According to yet another specific embodiment, the novel splice variantis an isolated protein or polypeptide having an amino acid sequence asset forth in AA424839_(—)1_P11 (SEQ ID NO:24), or a sequence homologousthere, i.e., which possesses at least 80, or 90% sequence identitytherewith. According to another related embodiment, the isolatedpolypeptide is at least 95, 96, 97, 98 or 99% homologous toAA424839_(—)1_P11 (SEQ ID NO:24).

It is another embodiment of the invention to provide molecules andisolated polypeptides comprising the soluble ectodomain (ECD) of theILDR1 proteins and fragments thereof as well as nucleic acid sequencesencoding said soluble ectodomain, as well as fragments thereof andconjugates and the use thereof as therapeutics including their use inimmunotherapy (promoting or inhibiting immune costimulation).

According to yet further embodiments the present invention providesdiscrete portions of the ILDR1 proteins including different portions ofthe extracellular domain corresponding to residues 24-162 of sequencesAA424839_P3 (SEQ ID NO:22) and AA424839_P5 (SEQ ID NO:21), correspondingto amino acid sequence depicted in SEQ ID NO:75, or residues 24-457 ofAA424839_P7 (SEQ ID NO:23), corresponding to amino acid sequencedepicted in SEQ ID NO:76, or residues 24-105 of AA424839_(—)1_P11 (SEQID NO:24), corresponding to amino acid sequence depicted in SEQ IDNO:296, or residues 50-160 of AA424839_(—)1_P3 (SEQ ID NO:22),corresponding to amino acid sequence depicted in SEQ ID NO:301, orvariants thereof possessing at least 80% sequence identity, morepreferably at least 90% sequence identity therewith and even morepreferably at least 95, 96, 97, 98 or 99% sequence identity therewith.

It is another embodiment of the invention to provide an isolated orpurified soluble protein or nucleic acid sequence having or encoding theextracellular domain of the ILDR1 protein which optionally may bedirectly or indirectly attached to a non-ILDR1 protein or nucleic acidsequence such as a soluble immunoglobulin domain or fragment.

According to certain embodiments, the present invention provides novelsplice variants of a known hypothetical protein LOC253012 isoform 1 (SEQID NO:35) (RefSeq accession identifier NP_(—)001034461) or apolynucleotide encoding same, and their use as diagnostic markers and/oras therapeutic agents which agonize or antagonize the binding of othermoieties to the LOC253012 proteins and/or which modulate (agonize orantagonize) at least one LOC253012 related biological activity.

According to one embodiment, the novel LOC253012 splice variant is anisolated polynucleotide comprising a nucleic acid having a nucleic acidsequence as set forth in any one of H68654_(—)1_T8 (SEQ ID NO:28),H68654_(—)1_T15 (SEQ ID NO:29), H68654_(—)1_T16 (SEQ ID NO:30),H68654_(—)1_T17 (SEQ ID NO:31), H68654_(—)1_T18 (SEQ ID NO:32),H68654_(—)1_T19 (SEQ ID NO:33), or H68654_(—)1_T20 (SEQ ID NO:34) or asequence homologous thereto. According to another embodiment, theisolated polynucleotide is at least 95% homologous to any one ofH68654_(—)1_T8 (SEQ ID NO:28), H68654_(—)1_T15 (SEQ ID NO:29),H68654_(—)1_T16 (SEQ ID NO:30), H68654_(—)1_T17 (SEQ ID NO:31),H68654_(—)1_T18 (SEQ ID NO:32), H68654_(—)1_T19 (SEQ ID NO:33), orH68654_(—)1_T20 (SEQ ID NO:34).

According to yet another embodiment, the novel LOC253012 splice variantis an isolated protein or polypeptide having an amino acid sequence asset forth in any one of H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12(SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ IDNO:40) or a sequence homologous thereto. According to anotherembodiment, the isolated polypeptide is at least 95, 96, 97, 98 or 99%homologous to any one of H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12(SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ IDNO:40).

It is another object of the invention to provide molecules and isolatedpolypeptides comprising the soluble ectodomain (ECD) of the LOC253012proteins and fragments thereof as well as nucleic acid sequencesencoding said soluble ectodomain, as well as fragments thereof andconjugates and the use thereof as therapeutics including their use inimmunotherapy (promoting or inhibiting immune costimulation).

According to yet further embodiments of the present invention there arediscrete portions of the LOC253012 proteins including different portionsof the extracellular domain corresponding to residues 38-349 of thesequence H68654_(—)1_P2 (SEQ ID NO:35), corresponding to amino acidsequence depicted in SEQ ID NO:144, or residues 19-337 of the sequencesH68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), corresponding to amino acid sequencedepicted in SEQ ID NO:145, or residues 1-335 of the sequencesH68654_(—)1_P5 (SEQ ID NO:36), corresponding to amino acid sequencedepicted in SEQ ID NO:300, or variants thereof possessing at least 80%sequence identity, more preferably at least 90% sequence identitytherewith and even more preferably at least 95, 96, 97, 98 or 99%sequence identity therewith.

It is another object of the invention to provide an isolated or purifiedsoluble protein or nucleic acid sequence encoding having or encoding theextracellular domain of the LOC253012 protein which optionally may bedirectly or indirectly attached to a non-LOC253012 protein or nucleicacid sequence such as a soluble immunoglobulin domain or fragment.

According to certain embodiments, the present invention provides novelsplice variants of AI216611, or a polynucleotide encoding same, whichcan be used as diagnostic markers and/or therapeutic agents whichagonize or antagonize the binding of other moieties to the AI216611proteins and/or which modulate (agonize or antagonize) at least oneAI216611 related biological activity.

According to one embodiment, the novel AI216611 splice variant is anisolated polynucleotide comprising a nucleic acid having a nucleic acidsequence as set forth in AI216611_T1 (SEQ ID NO:42), or a sequencehomologous thereto. According to another embodiment, the isolatedpolynucleotide is at least 95, 96, 97, 98 or 99% homologous toAI216611_T1 (SEQ ID NO:42).

According to yet another embodiment, the novel AI216611 splice variantis an isolated protein or polypeptide having an amino acid sequence asset forth in AI216611_P1 (SEQ ID NO:44) or a sequence homologousthereto. According to another embodiment, the isolated polypeptide is atleast 95, 96, 97, 98 or 99% homologous to AI216611_P1 (SEQ ID NO:44).

It is another object of the invention to provide molecules and isolatedpolypeptides comprising the soluble ectodomain (ECD) of the AI216611proteins and fragments thereof as well as nucleic acid sequencesencoding said soluble ectodomain, as well as fragments thereof andconjugates and the use thereof as therapeutics including their use inimmunotherapy (such as promoting or inhibiting immune costimulation).According to yet further embodiments of the present invention there arediscrete portions of the AI216611 proteins including different portionsof the extracellular domain corresponding to residues 29-147 of thesequence AI216611_P0 (SEQ ID NO:43) or AI216611_P1 (SEQ ID NO:44),corresponding to amino acid sequence depicted in SEQ ID NO:146, orresidues 1-145 of the sequence AI216611_P0 (SEQ ID NO:43), correspondingto amino acid sequence depicted in SEQ ID NO:298, or variants thereofpossessing at least 80% sequence identity, more preferably at least 90%sequence identity therewith and even more preferably at least 95, 96,97, 98 or 99% sequence identity therewith.

It is another object of the invention to provide an isolated or purifiedsoluble protein or nucleic acid sequence having or encoding theextracellular domain of the AI216611 protein which optionally may bedirectly or indirectly attached to a non-AI216611 protein or nucleicacid sequence such as a soluble immunoglobulin domain or fragment.

It is another object of the invention to provide vectors such asplasmids and recombinant viral vectors and host cells containing thatexpress AI216611, its secreted or soluble form and/or the ECD of theAI216611 protein and variants thereof or polypeptide conjugatescontaining any of the foregoing.

According to certain embodiments, the present invention provides novelsplice variants of a known hypothetical protein LOC387597 (SEQ ID NO:47)(RefSeq accession identifier NP_(—)955383, synonyms: NP_(—)955383;LISCH-like; C1ORF32; RP4-782G3.2; dJ782G3.1) or a polynucleotideencoding same, which can be used as diagnostic markers and/ortherapeutic agents which agonize or antagonize the binding of othermoieties to the C1ORF32 proteins and/or which modulate (agonize orantagonize) at least one C1ORF32 related biological activity.

According to one embodiment, the novel LOC387597 splice variant is anisolated polynucleotide comprising a nucleic acid having a nucleic acidsequence as set forth in any one of H19011_(—)1_T8 (SEQ ID NO:45),H19011_(—)1_T9 (SEQ ID NO:46), or a sequence homologous thereto.According to another embodiment, the isolated polynucleotide is at least95, 96, 97, 98 or 99% homologous to any one of H19011_(—)1_T8 (SEQ IDNO:45), H19011_(—)1_T9 (SEQ ID NO:46).

According to yet another embodiment, the novel splice LOC387597 variantis an isolated protein or polypeptide having an amino acid sequence asset forth in any one of H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9(SEQ ID NO:50) or a sequence homologous thereto. According to anotherembodiment, the isolated polypeptide is at least 95, 96, 97, 98 or 99%homologous to any one of H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9(SEQ ID NO:50).

It is another object of the invention to provide molecules and isolatedpolypeptides comprising the soluble ectodomain (ECD) of the C1ORF32proteins and fragments thereof as well as nucleic acid sequencesencoding said soluble ectodomain, as well as fragments thereof andconjugates and the use thereof as therapeutics including their use inimmunotherapy (promoting or inhibiting immune costimulation).

According to yet further embodiments of the present invention there arediscrete portions of the C1ORF32 proteins including different portionsof the extracellular domain corresponding to residues 21-186 of thesequence H19011_(—)1_P8 (SEQ ID NO:48), corresponding to amino acidsequence depicted in SEQ ID NO:147, or residues 21-169 of the sequenceH19011_(—)1_P9 (SEQ ID NO:50), corresponding to amino acid sequencedepicted in SEQ ID NO:148, or residues 1-184 of the sequenceH19011_(—)1_P8 (SEQ ID NO:48), corresponding to amino acid sequencedepicted in SEQ ID NO:299 or variants thereof possessing at least 80%sequence identity, more preferably at least 90% sequence identitytherewith and even more preferably at least 95, 96, 97, 98 or 99%sequence identity therewith.

It is another object of the invention to provide an isolated or purifiedsoluble protein or nucleic acid sequence encoding having or encoding theextracellular domain of the C1ORF32 protein which optionally may bedirectly or indirectly attached to a non-C1ORF32 protein or nucleic acidsequence such as a soluble immunoglobulin domain or fragment.

According to certain embodiments, the present invention provides novelsplice variants of known protein FXYD3, FXYD domain-containing iontransport regulator 3 precursor (SEQ ID NO:70) (SwissProt accessionidentifier FXYD3_HUMAN; known also according to the synonyms Chlorideconductance inducer protein Mat-8; Mammary tumor 8 kDa protein;Phospholemman-like) or a polynucleotide encoding same, which can be usedas diagnostic markers and/or therapeutic agents which agonize orantagonize the binding of other moieties to the FXYD3 proteins and/orwhich modulate (agonize or antagonize) at least one FXYD3 relatedbiological activity.

According to one embodiment, the novel FXYD3 splice variant is anisolated polynucleotide comprising a nucleic acid having a nucleic acidsequence as set forth in any one of R31375_T19 (SEQ ID NO:65),R31375_T25 (SEQ ID NO:66), R31375_T26 (SEQ ID NO:67), R31375_T29 (SEQ IDNO:68), R31375_T39 (SEQ ID NO:69), or a sequence homologous thereto.According to another embodiment, the isolated polynucleotide is at least95, 96, 97, 98 or 99% homologous to any one of R31375_T19 (SEQ IDNO:65), R31375_T25 (SEQ ID NO:66), R31375_T26 (SEQ ID NO:67), R31375_T29(SEQ ID NO:68), R31375_T39 (SEQ ID NO:69).

According to yet another embodiment, the novel FXYD3 splice variant isan isolated protein or polypeptide having an amino acid sequence as setforth in any one of R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ IDNO:73), R31375_P33 (SEQ ID NO:74) or a sequence homologous thereto.According to another embodiment, the isolated polypeptide is at least95, 96, 97, 98 or 99% homologous to any one of R31375_P14 (SEQ IDNO:72), R31375_P31 (SEQ ID NO:73), R31375_P33 (SEQ ID NO:74).

It is another object of the invention to provide molecules and isolatedpolypeptides comprising the soluble ectodomain (ECD) of the FXYD3proteins and fragments thereof as well as nucleic acid sequencesencoding said soluble ectodomain, as well as fragments thereof andconjugates and the use thereof as therapeutics including their use incancer immunotherapy.

According to yet further embodiments of the present invention there arediscrete portions of the FXYD3 proteins including different portions ofthe extracellular domain corresponding to residues 21-36 of the sequenceR31375_P0 (SEQ ID NO:70) or R31375_P31 (SEQ ID NO:73), corresponding toamino acid sequence depicted in SEQ ID NO:149, or residues 21-65 of thesequence R31375_P14 (SEQ ID NO:72), corresponding to amino acid sequencedepicted in SEQ ID NO:150, or residues 21-25 of the sequence R31375_P33(SEQ ID NO:74), corresponding to amino acid sequence depicted in SEQ IDNO:151, or residues 1-63 of the sequence R31375_P14 (SEQ ID NO:72),corresponding to amino acid sequence depicted in SEQ ID NO:297, orvariants thereof possessing at least 80% sequence identity, morepreferably at least 90% sequence identity therewith and even morepreferably at least 95, 96, 97, 98 or 99% sequence identity therewith.

It is another object of the invention to provide an isolated or purifiedsoluble protein or nucleic acid sequence encoding having or encoding theextracellular domain of any one of the VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 or FXYD3 proteins which optionally may be directly orindirectly attached to a non-VSIG1, non-ILDR1, non-LOC253012,non-AI216611, non-C1ORF32 or non-FXYD3 protein or nucleic acid sequence,respectively, such as a soluble immunoglobulin domain or fragment.

It is another object of the invention to provide molecules and isolatedpolypeptides comprising edge portion, tail or head portion, of any oneof the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 novel variantsof the invention, or a homologue or a fragment thereof as well asnucleic acid sequences encoding said edge portion, tail or head portion,as well as fragments thereof and conjugates and the use thereof astherapeutics and/or for diagnostics.

It is further object of the invention to provide molecules and isolatedpolypeptides comprising a bridge, edge portion, tail or head portion, asdepicted in any one of SEQ. ID NOs: 284-295, or a homologue or afragment thereof as well as nucleic acid sequences encoding said edgeportion, tail or head portion, as well as fragments thereof andconjugates and the use thereof as therapeutics and/or for diagnostics.

It is another object of the invention to provide vectors such asplasmids and recombinant viral vectors and host cells containing thevectors that express any one of VSIG1, ILDR1, LOC253012, AI216611,C1ORF32, FXYD3, its secreted or soluble form and/or the ECD of theVSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 protein and variantsthereof or polypeptide conjugates containing any of the foregoing.

It is another object of the invention to use these vectors such asplasmids and recombinant viral vectors and host cells containing thatexpress any one of VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3,its secreted or soluble form and/or the ECD of the VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, FXYD3 protein and variants thereof orpolypeptide conjugates containing any of the foregoing to produce saidVSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 protein, fragments orvariants thereof and/or conjugates containing any one of the foregoing.

It is another object of the invention to provide pharmaceutical ordiagnostic compositions containing any of the foregoing.

It is another object of the invention to provide and use compoundsincluding VSIG1 ectodomain or fragments or variants thereof, which aresuitable for treatment or prevention of cancer, autoimmune disorders,transplant rejection, graft versus host disease, and/or for blocking orpromoting immune costimulation mediated by the VSIG1, ILDR1, LOC253012,AI216611, FXYD3 or C1ORF32 polypeptide.

It is a specific object of the invention to develop novel monoclonal orpolyclonal antibodies and antibody fragments and conjugates containingthat specifically bind the full length VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 or FXYD3 antigen, selected from the group consistingof AI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5(SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15),AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5(SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23), AA424839_(—)1_P11 (SEQ IDNO:24), H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73), R31375_P33 (SEQ IDNO:74), its secreted form and/or the ECD thereof or conjugates orfragments thereof. These antibodies are potentially useful astherapeutics and/or diagnostic agents (both in vitro and in vivodiagnostic methods). Included in particular are antibodies and fragmentsthat are immune activating or immune suppressing such as antibodies orfragments that target cells via ADCC (antibody dependent cellularcytotoxicity) or CDC (complement dependent cytotoxicity) activities.

It is another object of the invention to provide diagnostic methods thatinclude the use of any of the foregoing including by way of exampleimmunohistochemical assay, radioimaging assays, in-vivo imaging,radioimmunoassay (RIA), ELISA, slot blot, competitive binding assays,fluorimetric imaging assays, Western blot, FACS, and the like. Inparticular this includes assays which use chimeric or non-humanantibodies or fragments that specifically bind the intact VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 protein, selected from the groupconsisting of AI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12),AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQID NO:15), AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22),AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23),AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ ID NO:35),H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73), R31375_P33 (SEQ ID NO:74), its soluble form, its ECD,and or conjugates, fragments or variants thereof.

It is another object of the invention to use novel therapeuticallyeffective polyclonal or monoclonal antibodies against anyone of theVSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 antigen, selectedfrom the group consisting of AI581519_P3 (SEQ ID NO:11), AI581519_P4(SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14),AI581519_P9 (SEQ ID NO:15), AI581519_P10 (SEQ ID NO:16), AA424839_P3(SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23),AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ ID NO:35),H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73), R31375_P33 (SEQ ID NO:74), and fragments, conjugates,and variants thereof for treating conditions wherein the VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 antigen or its secreted or solubleform or ECD and/or portions or variants thereof are differentiallyexpressed including various cancers and malignancies including non-solidand solid tumors, sarcomas, hematological malignancies including but notlimited to acute lymphocytic leukemia, chronic lymphocytic leukemia,acute myelogenous leukemia, chronic myelogenous leukemia, multiplemyeloma, Hodgkin's lymphoma, Non-Hodgkin's lymphoma, cancer of thebreast, prostate, lung, ovary, colon, spleen, kidney, bladder, head andneck, uterus, testicles, stomach, cervix, liver, bone, skin, pancreas,brain and wherein the cancer is non-metastatic, invasive or metastatic.

It is another object of the invention to use novel therapeuticallyeffective polyclonal or monoclonal antibodies against anyone of theVSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 antigen, selectedfrom the group consisting of AI581519_P3 (SEQ ID NO:11), AI581519_P4(SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14),AI581519_P9 (SEQ ID NO:15), AI581519_P10 (SEQ ID NO:16), AA424839_P3(SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23),AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ ID NO:35),H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73), R31375_P33 (SEQ ID NO:74), and fragments, conjugates andvariants thereof for treating non-malignant disorders such as immunedisorders including but not limited to autoimmune diseases, transplantrejection and graft versus host disease.

It is a specific object of the invention to use antibodies and antibodyfragments against VSIG1 antigen, its secreted or soluble form or ECDand/or variants, conjugates, or fragments thereof and fragments andvariants thereof for treating and diagnosing lung cancer and/or ovariancancer, wherein this antigen is differentially expressed.

It is a specific embodiment of the invention to use antibodies andantibody fragments against ILDR1 antigen, its secreted or soluble formor ECD and/or variants, conjugates, or fragments thereof and fragmentsand variants thereof for treating and diagnosing colon and/or ovariancancers wherein this antigen is differentially expressed.

It is a specific object of the invention to use antibodies and antibodyfragments against LOC253012 or C1ORF32 antigen, its secreted or solubleform or ECD and/or variants, conjugates, or fragments thereof andfragments and variants thereof for treating and diagnosing lung cancer,particularly small cell lung carcinoma, wherein this antigen isdifferentially expressed.

It is a specific object of the invention to use antibodies and antibodyfragments against AI216611 antigen, its secreted or soluble form or ECDand/or variants, conjugates, or fragments thereof and fragments andvariants thereof for treating and diagnosing colon cancer, wherein thisantigen is differentially expressed.

It is a specific object of the invention to use antibodies and antibodyfragments against FXYD3 wild type antigen (R31375_P0 (SEQ ID NO:70)), orantibodies and antibody fragments against its secreted or soluble formor ECD and conjugates containing for treating and diagnosing ovariancancer, wherein this antigen is differentially expressed.

It is another object of the invention to use antibodies and antibodyfragments, and conjugates containing, against the VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 antigen, selected from the groupconsisting of AI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12),AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQID NO:15), AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22),AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23),AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ ID NO:35),H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73), R31375_P33 (SEQ ID NO:74) in modulating (enhancing orinhibiting) immunity including antibodies that activate or suppress theimmune co-stimulation in particular B7 related immune costimulation andare capable of treating related therapeutic applications, throughpositive stimulation of T cell activity against cancer cells, andnegative stimulation of T cell activity for the treatment ofautoimmunity and other immune disorders.

It is another specific object of the invention to produce antibodies andantibody fragments against discrete portions of the VSIG1 proteinsincluding different portions of the extracellular domain correspondingto residues 23-234 of the VSIG1 protein sequence contained in thesequence of AI581519_P3 (SEQ ID NO:11) corresponding to amino acidsequence depicted in SEQ ID NO:138, or residues 23-270 of the VSIG1protein sequence contained in the sequence of AI581519_P4 (SEQ ID NO:12)corresponding to amino acid sequence depicted in SEQ ID NO:139, orresidues 23-296 of the VSIG1 protein sequence contained in the sequenceof AI581519_P5 (SEQ ID NO:13) corresponding to amino acid sequencedepicted in SEQ ID NO:140, or residues 23-193 of the VSIG1 proteinsequence contained in the sequence of AI581519_P7 (SEQ ID NO:14)corresponding to amino ac, id sequence depicted in SEQ ID NO:141, orresidues 23-203 of the VSIG1 protein sequence contained in the sequenceof AI581519_P9 (SEQ ID NO:15) corresponding to amino acid sequencedepicted in SEQ ID NO:142, or residues 23-231 of the VSIG1 proteinsequence contained in the sequence of AI581519_P10 (SEQ ID NO:16),corresponding to amino acid sequence depicted in SEQ ID NO:143, orresidues 26-293 of the VSIG1 protein sequence contained in the sequenceof AI581519_P5 (SEQ ID NO:13), corresponding to amino acid sequencedepicted in SEQ ID NO:302.

It is another specific embodiment of the invention to produce antibodiesand antibody fragments against discrete portions of the ILDR1 proteinsincluding different portions of the extracellular domain correspondingto residues 24-162 of sequences AA424839_P3 (SEQ ID NO:22) andAA424839_P5 (SEQ ID NO:21), corresponding to amino acid sequencedepicted in SEQ ID NO:75, residues 24-457 of AA424839_P7 (SEQ ID NO:23),corresponding to amino acid sequence depicted in SEQ ID NO:76, andresidues 24-105 of AA424839_(—)1_P11 (SEQ ID NO:24), corresponding toamino acid sequence depicted in SEQ ID NO:296, or residues 50-160 ofAA424839_(—)1_P3 (SEQ ID NO:22), corresponding to amino acid sequencedepicted in SEQ ID NO:301 of the ILDR1 protein sequences disclosedherein.

It is another specific object of the invention to produce antibodies andantibody fragments against discrete portions of the LOC253012 proteinsincluding different portions of the extracellular domain correspondingto residues 38-349 of the sequence H68654_(—)1_P2 (SEQ ID NO:35),corresponding to amino acid sequence depicted in SEQ ID NO:144, orresidues 19-337 of the sequences H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40), orresidues 1-335 of the sequences H68654_(—)1_P5 (SEQ ID NO:36),corresponding to amino acid sequence depicted in SEQ ID NO:300, of theLOC253012 protein sequences disclosed herein.

It is another specific object of the invention to produce antibodies andantibody fragments against discrete portions of the AI216611 proteinsincluding different portions of the extracellular domain correspondingto residues 29-147 of the sequence AI216611_P0 (SEQ ID NO:43) orAI216611_P1 (SEQ ID NO:44), corresponding to amino acid sequencedepicted in SEQ ID NO:146, or residues 1-145 of the sequence AI216611_P0(SEQ ID NO:43), corresponding to amino acid sequence depicted in SEQ IDNO:298 sequence disclosed herein.

It is another specific object of the invention to produce antibodies andantibody fragments against discrete portions of the C1ORF32 proteinsincluding different portions of the extracellular domain correspondingto residues 21-186 of the C1ORF32 protein sequence contained in thesequence of H19011_(—)1_P8 (SEQ ID NO:48), corresponding to amino acidsequence depicted in SEQ ID NO:147, or residues 21-169 of the C1ORF32protein sequence contained in the sequence of H19011_(—)1_P9 (SEQ IDNO:50), corresponding to amino acid sequence depicted in SEQ ID NO:148,or residues 1-184 of the sequence H19011_(—)1_P8 (SEQ ID NO:48),corresponding to amino acid sequence depicted in SEQ ID NO:299.

It is another specific object of the invention to produce antibodies andantibody fragments against discrete portions of the FXYD3 proteinsincluding different portions of the extracellular domain correspondingto residues 21-36 of the FXYD3 protein sequence contained in thesequence of R31375_P0 (SEQ ID NO:70) or R31375_P31 (SEQ ID NO:73),corresponding to amino acid sequence depicted in SEQ ID NO:149, orresidues 21-65 of the FXYD3 protein sequence contained in the sequenceof R31375_P14 (SEQ ID NO:72), corresponding to amino acid sequencedepicted in SEQ ID NO:150, or residues 21-25 of the FXYD3 proteinsequence contained in the sequence of R31375_P33 (SEQ ID NO:74),corresponding to amino acid sequence depicted in SEQ ID NO:151, orresidues 1-63 of the sequence R31375_P14 (SEQ ID NO:72), correspondingto amino acid sequence depicted in SEQ ID NO:297.

It is a specific object of the invention to provide polyclonal andmonoclonal antibodies and fragments thereof or an antigen bindingfragment thereof comprising an antigen bindings site that bindsspecifically to the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3proteins, its soluble forms, the ECD thereof and/or variants andfragments thereof.

It is a specific object of the invention to use such antibodies andfragments thereof for treatment or prevention of cancer and/or formodulating (activating or blocking) the activity of the target in theimmune co-stimulatory system.

It is a related object of the invention to select monoclonal andpolyclonal antibodies and fragments thereof against VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 which are suitable for treatmentor prevention of autoimmune disorders, transplant rejection, GVHD,and/or for blocking or enhancing immune costimulation mediated by theVSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 polypeptide.

It is a specific object of the invention to use antibodies againstanyone of the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3antigen, soluble form, ECD or fragment or variant thereof for thetreatment and diagnosis of cancers including by way of example lungcancer, ovarian cancer, colon cancer, as well as other non-solid andsolid tumors, sarcomas, hematological malignancies including but notlimited to acute lymphocytic leukemia, chronic lymphocytic leukemia,acute myelogenous leukemia, chronic myelogenous leukemia, multiplemyeloma, Hodgkin's lymphoma, Non-Hodgkin's lymphoma, cancer of thebreast, prostate, spleen, kidney, bladder, head and neck, uterus,testicles, stomach, cervix, liver, bone, skin, pancreas, brain andwherein the cancer is non-metastatic, invasive or metastatic.

With regard to lung cancer, the disease is selected from the groupconsisting of squamous cell lung carcinoma, lung adenocarcinoma,carcinoid, small cell lung cancer or non-small cell lung cancer.

It is another object of the invention to provide and use antibodies andantibody fragments against anyone of the VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 or FXYD3 antigen, its soluble form, or ECD andvariants or fragments thereof as well as soluble polypeptides containingthe ectodomain of the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 orFXYD3 antigen or a portion thereof which are useful for immunemodulation, including treatment of autoimmunity and preferably fortreating an autoimmune disease selected from autoimmune diseases:Multiple sclerosis; Psoriasis; Rheumatoid arthritis; Systemic lupuserythematosus; Ulcerative colitis; Crohn's disease; immune disordersassociated with graft transplantation rejection, benign lymphocyticangiitis, lupus erythematosus, Hashimoto's thyroiditis, primarymyxedema, Graves' disease, pernicious anemia, autoimmune atrophicgastritis, Addison's disease, insulin dependent diabetes mellitus, goodpasture's syndrome, myasthenia gravis, pemphigus, sympatheticophthalmia, autoimmune uveitis, autoimmune hemolytic anemia, idiopathicthrombocytopenia, primary biliary cirrhosis, chronic action hepatitis,ulceratis colitis, Sjogren's syndrome, rheumatic disease, polymyositis,scleroderma, mixed connective tissue disease, inflammatory rheumatism,degenerative rheumatism, extra-articular rheumatism, collagen diseases,chronic polyarthritis, psoriasis arthropathica, ankylosing spondylitis,juvenile rheumatoid arthritis, periarthritis humeroscapularis,panarteriitis nodosa, progressive systemic scleroderma, arthritisuratica, dermatomyositis, muscular rheumatism, myositis, myogelosis andchondrocalcinosis.

It is another object of the invention to provide and use compoundsincluding drugs such as small molecules, peptides, antibodies andfragments that bind anyone of the VSIG1, ILDR1, LOC253012, AI216611,C1ORF32 or FXYD3 antigen, as well as ribozymes or antisense or siRNAswhich target the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3nucleic acid sequence or fragments or variants thereof which are usefulfor treatment or prevention of cancer, autoimmune disorders, transplantrejection, GVHD, and/or for blocking or enhancing immune costimulationmediated by the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3polypeptide.

It is another object of the invention to provide and use compoundsincluding drugs such as small molecules, peptides, antibodies andfragments that bind the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 orFXYD3 antigen, as well as ribozymes or antisense or siRNAs which targetthe FXYD3 nucleic acid sequence or fragments or variants thereof whichare useful for treatment or prevention of cancer.

It is a preferred object to provide therapeutic and diagnosticantibodies and fragments and conjugates containing useful in treating ordiagnosing any of the foregoing that specifically bind to amino-acidsresidues 23-234 of the sequence AI581519_P3 (SEQ ID NO:11),corresponding to amino acid sequence depicted in SEQ ID NO:138, orresidues 23-270 of the sequence AI581519_P4 (SEQ ID NO:12),corresponding to amino acid sequence depicted in SEQ ID NO:139, orresidues 23-296 of the sequence AI581519_P5 (SEQ ID NO:13),corresponding to amino acid sequence depicted in SEQ ID NO:140, orresidues 23-193 of the sequence AI581519_P7 (SEQ ID NO:14),corresponding to amino acid sequence depicted in SEQ ID NO:141, orresidues 23-203 of the sequence AI581519_P9 (SEQ ID NO:15),corresponding to amino acid sequence depicted in SEQ ID NO:144, orresidues 23-231 of the sequence AI581519_P10 (SEQ ID NO:16),corresponding to amino acid sequence depicted in SEQ ID NO:143, orresidues 26-293 of the sequence AI581519_P5 (SEQ ID NO:13),corresponding to amino acid sequence depicted in SEQ ID NO:302 of theVSIG1 protein sequences disclosed herein.

It is a preferred embodiment to provide therapeutic and diagnosticantibodies and fragments and conjugates containing useful in treating ordiagnosing any of the foregoing that specifically bind to amino-acidsresidues 24-162 of the ILDR1 protein sequence contained in the sequenceof AA424839_P3 (SEQ ID NO:22) and AA424839_P5 (SEQ ID NO:21),corresponding to amino acid sequence depicted in SEQ ID NO:75, residues24-457 of the ILDR1 protein sequence contained in the sequence ofAA424839_P7 (SEQ ID NO:23), corresponding to amino acid sequencedepicted in SEQ ID NO:76, and residues 24-105 of the ILDR1 proteinsequence contained in the sequence of AA424839_(—)1_P11 (SEQ ID NO:24),corresponding to amino acid sequence depicted in SEQ ID NO:296, orresidues 50-160 of AA424839_(—)1_P3 (SEQ ID NO:22), corresponding toamino acid sequence depicted in SEQ ID NO:301.

It is a preferred object to provide therapeutic and diagnosticantibodies and fragments and conjugates containing useful in treating ordiagnosing any of the foregoing that specifically bind to amino-acidsresidues 38-349 of the LOC253012 protein sequence contained in thesequence of H68654_(—)1_P2 (SEQ ID NO:35), corresponding to amino acidsequence depicted in SEQ ID NO:144, or residues 19-337 of the of theLOC253012 protein sequence contained in the sequences of H68654_(—)1_P5(SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ IDNO:38), H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40)),corresponding to amino acid sequence depicted in SEQ ID NO:145, orresidues 1-335 of the sequences H68654_(—)1_P5 (SEQ ID NO:36),corresponding to amino acid sequence depicted in SEQ ID NO:300.

It is a preferred object to provide therapeutic and diagnosticantibodies and fragments and conjugates containing useful in treating ordiagnosing any of the foregoing that specifically bind to amino-acidsresidues 29-147 of the AI216611 protein sequence contained in thesequence of AI216611_P0 (SEQ ID NO:43) or AI216611_P1 (SEQ ID NO:44),corresponding to amino acid sequence depicted in SEQ ID NO:146, orresidues 1-145 of the sequence AI216611_P0 (SEQ ID NO:43), correspondingto amino acid sequence depicted in SEQ ID NO:298.

It is a preferred object to provide therapeutic and diagnosticantibodies and fragments and conjugates containing useful in treating ordiagnosing any of the foregoing that specifically bind to amino-acidsresidues 21-186 of the C1ORF32 protein sequence contained in thesequence of H19011_(—)1_P8 (SEQ ID NO:48), corresponding to amino acidsequence depicted in SEQ ID NO:147, or residues 21-169 of the sequenceof the C1ORF32 protein sequence contained in the sequence ofH19011_(—)1_P9 (SEQ ID NO:50), corresponding to amino acid sequencedepicted in SEQ ID NO:149, or residues 1-184 of the sequenceH19011_(—)1_P8 (SEQ ID NO:48), corresponding to amino acid sequencedepicted in SEQ ID NO:299.

It is a preferred object to provide therapeutic and diagnosticantibodies and fragments and conjugates containing useful in treating ordiagnosing any of the foregoing that specifically bind to amino-acidsresidues 21-36 of the FXYD3 protein sequence contained in the sequenceof R31375_P0 (SEQ ID NO:70), or R31375_P31 (SEQ ID NO:73), correspondingto amino acid sequence depicted in SEQ ID NO:149 or residues 21-65 ofthe FXYD3 protein sequence contained in the sequence of R31375_P14 (SEQID NO:72), corresponding to amino acid sequence depicted in SEQ IDNO:150, or residues or residues 21-25 of the FXYD3 protein sequencecontained in the sequence of R31375_P33 (SEQ ID NO:74), corresponding toamino acid sequence depicted in SEQ ID NO:151, or residues 1-63 of thesequence R31375_P14 (SEQ ID NO:72), corresponding to amino acid sequencedepicted in SEQ ID NO:297.

It is also a preferred object to provide antibodies and fragmentsthereof that bind to VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3and the specific residues above-identified and fragments thereof,wherein the antibody is a chimeric, humanized, fully human antibodyand/or is an antibody or antibody fragment having CDC or ADCC activitieson target cells.

It is also a preferred object to provide chimeric and human antibodiesand fragments thereof and conjugates containing that bind to VSIG1,ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 and the specific residuesabove-identified and fragments thereof.

It is another specific object of the invention to provide antibodyfragments and conjugates containing useful in the foregoing therapiesand related diagnostic methods including but not limited to Fab,F(ab′)2, Fv or scFv fragment.

It is also an object of the invention to directly or indirectly attachthe subject antibodies and fragments to markers and other effectormoieties such as a detectable marker, or to an effector moiety such asan enzyme, a toxin, a therapeutic agent, or a chemotherapeutic agent.

In a preferred embodiment the inventive antibodies or fragments may beattached directly or indirectly to a radioisotope, a metal chelator, anenzyme, a fluorescent compound, a bioluminescent compound or achemiluminescent compound.

It is also an object of the invention to provide pharmaceutical anddiagnostic compositions that comprise a therapeutically ordiagnostically effective form of an antibody or antibody fragmentaccording to the invention.

It is another specific object of the invention to inhibit the growth ofcells that express VSIG1 in a subject, comprising: administering to saidsubject an antibody that specifically binds to the antigen referred toherein as AI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12),AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQID NO:15), AI581519_P10 (SEQ ID NO:16) or VSIG1.

It is another specific object of the invention to provide methods fortreating or preventing cancer, comprising administering to a patient aneffective amount of a monoclonal antibody that specifically bindAI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15),AI581519_P10 (SEQ ID NO:16) or VSIG1.

It is a more preferred object of the invention to use these antibodiesfor treating cancers selected from the group consisting of lung cancer,and ovarian cancer, and wherein the lung cancer or the ovarian cancer isnon-metastatic, invasive or metastatic, wherein preferably the antibodyhas an antigen-binding region specific for the extracellular domain ofAI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15),AI581519_P10 (SEQ ID NO:16).

It is another object of the invention to provide methods for treating orpreventing autoimmune diseases, comprising administering to a patient aneffective amount of a polyclonal or monoclonal antibody or fragment or aconjugate containing that specifically bind AI581519_P3 (SEQ ID NO:11),AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQID NO:14), AI581519_P9 (SEQ ID NO:15), AI581519_P10 (SEQ ID NO:16).

It is another specific embodiment of the invention to inhibit the growthof cells that express ILDR1 in a subject, comprising: administering tosaid subject an antibody that specifically binds to the antigen referredto herein as AA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21),AA424839_P7 (SEQ ID NO:23), AA424839_(—)1_P11 (SEQ ID NO:24) or ILDR1.

It is another specific embodiment of the invention to provide methodsfor treating or preventing cancer, comprising administering to a patientan effective amount of a monoclonal antibody that specifically binds toAA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQID NO:23), AA424839_(—)1_P11 (SEQ ID NO:24) or ILDR1.

It is a more preferred embodiment of the invention to use theseantibodies for treating cancers selected from the group consisting ofcolon cancer or ovarian cancer, and wherein the colon cancer or theovarian cancer is non-metastatic, invasive or metastatic whereinpreferably the antibody has an antigen-binding region specific for theextracellular domain of AA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ IDNO:21), AA424839_P7 (SEQ ID NO:23) or AA424839_(—)1_P11 (SEQ ID NO:24).

It is another embodiment of the invention to provide methods fortreating or preventing autoimmune diseases, comprising administering toa patient an effective amount of a polyclonal or monoclonal antibody orfragment that specifically binds AA424839_P3 (SEQ ID NO:22), AA424839_P5(SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23), or AA424839_(—)1_P11 (SEQ IDNO:24).

It is another specific object of the invention to inhibit the growth ofcells that express LOC253012 in a subject, comprising: administering tosaid subject an antibody that specifically binds to the antigen referredto herein as H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ IDNO:36), H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40) orLOC253012.

It is another specific object of the invention to provide methods fortreating or preventing cancer, comprising administering to a patient aneffective amount of a monoclonal antibody that specifically bindH68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40) orLOC253012.

It is a more preferred object of the invention to use these antibodiesfor treating cancers selected from the group consisting of lung cancer,especially small cell lung carcinoma, and wherein the lung cancer isnon-metastatic, invasive or metastatic wherein preferably the antibodyhas an antigen-binding region specific for the extracellular domain ofH68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40).

It is another object of the invention to provide methods for treating orpreventing autoimmune diseases, comprising administering to a patient aneffective amount of a polyclonal or monoclonal antibody or fragment thatspecifically bind H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ IDNO:36), H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40).

It is another specific object of the invention to provide methods fortreating or preventing cancer, comprising administering to a patient aneffective amount of a monoclonal antibody that specifically binds toAI216611_P0 (SEQ ID NO:43) or AI216611_P1 (SEQ ID NO:44).

It is another object of the invention to provide methods for treating orpreventing autoimmune diseases, comprising administering to a patient aneffective amount of a polyclonal or monoclonal antibody or fragment or aconjugate containing that specifically bind AI216611_P0 (SEQ ID NO:43)or AI216611_P1 (SEQ ID NO:44).

It is another specific object of the invention to inhibit the growth ofcells that express C1ORF32 in a subject, comprising: administering tosaid subject an antibody that specifically binds to the antigen referredto herein as H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), or C1ORF32.

It is another specific object of the invention to provide methods fortreating or preventing cancer, comprising administering to a patient aneffective amount of a monoclonal antibody that specifically binds toH19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50) or C1ORF32.

It is a more preferred object of the invention to use these antibodiesfor treating cancers selected from the group consisting of lung cancer,particularly lung small cell carcinoma, and wherein the lung cancer isnon-metastatic, invasive or metastatic, wherein preferably the antibodyhas an antigen-binding region specific for the extracellular domain ofH19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50).

It is another object of the invention to provide methods for treating orpreventing autoimmune diseases, comprising administering to a patient aneffective amount of a polyclonal or monoclonal antibody or fragment thatspecifically bind H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50).

It is another specific object of the invention to inhibit the growth ofcells that express FXYD3 in a subject, comprising: administering to saidsubject an antibody that specifically binds to the antigen referred toherein as R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72),R31375_P31 (SEQ ID NO:73), R31375_P33 (SEQ ID NO:74).

It is another specific object of the invention to use part or all of theectodomain of VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 or itsvariants and conjugates containing for administration as an anti-cancervaccine, for immunotherapy of cancer, including but not limited toovarian cancer.

It is another specific object of the invention to provide methods fortreating or preventing cancer, comprising administering to a patient aneffective amount of a monoclonal antibody that specifically binds toR31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ IDNO:73), R31375_P33 (SEQ ID NO:74).

It is a more preferred object of the invention to use these antibodiesfor treating ovarian cancer, and wherein the ovarian cancer isnon-metastatic, invasive or metastatic, wherein preferably the antibodyhas an antigen-binding region specific for the extracellular domain ofR31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ IDNO:73), R31375_P33 (SEQ ID NO:74).

In another embodiment of the invention the cancer is selected from thegroup consisting of non-solid and solid tumors, sarcomas, hematologicalmalignancies including but not limited to acute lymphocytic leukemia,chronic lymphocytic leukemia, acute myelogenous leukemia, chronicmyelogenous leukemia, multiple myeloma, Hodgkin's lymphoma,Non-Hodgkin's lymphoma, cancer of the lung, ovary, breast, prostate,colon, spleen, kidney, bladder, head and neck, uterus, testicles,stomach, cervix, liver, bone, skin, pancreas, brain and wherein thecancer may be non-metastatic, invasive or metastatic.

In a preferred embodiment the autoimmune diseases include Multiplesclerosis; Psoriasis; Rheumatoid arthritis; Systemic lupuserythematosus; Ulcerative colitis; Crohn's disease; immune disordersassociated with graft transplantation rejection, benign lymphocyticangiitis, lupus erythematosus, Hashimoto's thyroiditis, primarymyxedema, Graves' disease, pernicious anemia, autoimmune atrophicgastritis, Addison's disease, insulin dependent diabetes mellitis, goodpasture's syndrome, myasthenia gravis, pemphigus, sympatheticophthalmia, autoimmune uveitis, autoimmune hemolytic anemia, idiopathicthrombocytopenia, primary biliary cirrhosis, chronic action hepatitis,ulceratis colitis, Sjogren's syndrome, rheumatic disease, polymyositis,scleroderma, mixed connective tissue disease, inflammatory rheumatism,degenerative rheumatism, extra-articular rheumatism, collagen diseases,chronic polyarthritis, psoriasis arthropathica, ankylosing spondylitis,juvenile rheumatoid arthritis, periarthritis humeroscapularis,panarteriitis nodosa, progressive systemic scleroderma, arthritisuratica, dermatomyositis, muscular rheumatism, myositis, myogelosis andchondrocalcinosis.

It is a specific object of the invention to provide methods for treatingor preventing rejection of any organ transplant and/or graft versus hostdisease, comprising administering to a patient an effective amount of anantibody that specifically bind AI581519_P3 (SEQ ID NO:11), AI581519_P4(SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14),AI581519_P9 (SEQ ID NO:15), AI581519_P10 (SEQ ID NO:16), AA424839_P3(SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23),AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ ID NO:35),H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73), R31375_P33 (SEQ ID NO:74). It is also preferred in theforegoing methods that the antibody possess an antigen-binding regionspecific for the extracellular domain of AI581519_P3 (SEQ ID NO:11),AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQID NO:14), AI581519_P9 (SEQ ID NO:15), AI581519_P10 (SEQ ID NO:16),AA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQID NO:23), or AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ IDNO:35), H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73), R31375_P33 (SEQ ID NO:74).

According to the present invention, each one of the following: the VSIG1ectodomain, ILDR1 ectodomain, LOC253012 ectodomain, AI216611 ectodomain,C1ORF32 ectodomain or FXYD3 ectodomain of the present invention,antibodies and fragments that bind the VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 of FXYD3 antigen, the compounds including drugs suchas small molecules, peptides, as well as ribozymes or antisense orsiRNAs which target the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 orFXYD3 nucleic acid sequence or fragments or variants thereof which areuseful for treatment or prevention of cancer, autoimmune disorders,transplant rejection, GVHD, and/or for blocking or enhancing immuneco-stimulation mediated by the VSIG1, ILDR1, LOC253012, AI216611,C1ORF32 or FXYD3 polypeptide, can be used with simultaneous blockade ofseveral co-stimulatory pathways or in combination therapy withconventional drugs, such as immunosuppressants or cytotoxic drugs forcancer.

It is another object of the invention to provide assays for detectingthe presence of AI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12),AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQID NO:15), AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22),AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23), orAA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ ID NO:35),H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73) or R31375_P33 (SEQ ID NO:74) protein in vitro or in vivoin a biological sample or individual comprising contacting the samplewith an antibody having specificity for AI581519_P3 (SEQ ID NO:11),AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQID NO:14), AI581519_P9 (SEQ ID NO:15), AI581519_P10 (SEQ ID NO:16),AA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQID NO:23), or AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ IDNO:35), H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73) or R31375_P33 (SEQ ID NO:74) polypeptides, or acombination thereof, and detecting the binding of AI581519_P3 (SEQ IDNO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13),AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15), AI581519_P10(SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21),AA424839_P7 (SEQ ID NO:23), or AA424839_(—)1_P11 (SEQ ID NO:24),H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73) or R31375_P33 (SEQID NO:74) protein in the sample.

It is another object of the invention to provide methods for detecting adisease, diagnosing a disease, monitoring disease progression ortreatment efficacy or relapse of a disease, or selecting a therapy for adisease, comprising detecting expression of a AI581519_P3 (SEQ IDNO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13),AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15), AI581519_P10(SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21),AA424839_P7 (SEQ ID NO:23), or AA424839_(—)1_P11 (SEQ ID NO:24),H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73) or R31375_P33 (SEQID NO:74).

In a related object the detected diseases will include cancers such aslung cancer, ovarian cancer, colon cancer, as well as other non-solidand solid tumors, sarcomas, hematological malignancies including but notlimited to acute lymphocytic leukemia, chronic lymphocytic leukemia,acute myelogenous leukemia, chronic myelogenous leukemia, multiplemyeloma, Hodgkin's lymphoma, Non-Hodgkin's lymphoma, cancer of thebreast, prostate, spleen, kidney, bladder, head and neck, uterus,testicles, stomach, cervix, liver, bone, skin, pancreas, brain andwherein the cancer is non-metastatic, invasive or metastatic.

With regard to lung cancer, the disease is selected from the groupconsisting of non-metastatic, invasive or metastatic lung cancer;squamous cell lung carcinoma, lung adenocarcinoma, carcinoid, small celllung cancer or non-small cell lung cancer; detection of overexpressionin lung metastasis (vs. primary tumor); detection of overexpression inlung cancer, for example non small cell lung cancer, for exampleadenocarcinoma, squamous cell cancer or carcinoid, or large cellcarcinoma; identification of a metastasis of unknown origin whichoriginated from a primary lung cancer; assessment of a malignant tissueresiding in the lung that is from a non-lung origin, including but notlimited to: osteogenic and soft tissue sarcomas; colorectal, uterine,cervix and corpus tumors; head and neck, breast, testis and salivarygland cancers; melanoma; and bladder and kidney tumors; distinguishingbetween different types of lung cancer, therefore potentially affectingtreatment choice (e.g. small cell vs. non small cell tumors); analysisof unexplained dyspnea and/or chronic cough and/or hemoptysis;differential diagnosis of the origin of a pleural effusion; diagnosis ofconditions which have similar symptoms, signs and complications as lungcancer and where the differential diagnosis between them and lung canceris of clinical importance including but not limited to: non-malignantcauses of lung symptoms and signs, including but not limited to: lunglesions and infiltrates, wheeze, stridor, tracheal obstruction,esophageal compression, dysphagia, recurrent laryngeal nerve paralysis,hoarseness, phrenic nerve paralysis with elevation of the hemidiaphragmand Horner syndrome; or detecting a cause of any condition suggestive ofa malignant tumor including but not limited to anorexia, cachexia,weight loss, fever, hypercalcemia, hypophosphatemia, hyponatremia,syndrome of inappropriate secretion of antidiuretic hormone, elevatedANP, elevated ACTH, hypokalemia, clubbing, neurologic-myopathicsyndromes and thrombophlebitis.

With regard to ovarian cancer, the compounds of the present inventioncan be used in the diagnosis, treatment or prognostic assessment ofnon-metastatic, invasive or metastatic ovarian cancer; correlating stageand malignant potential; identification of a metastasis of unknownorigin which originated from a primary ovarian cancer; differentialdiagnosis between benign and malignant ovarian cysts; diagnosing a causeof infertility, for example differential diagnosis of various causesthereof; detecting of one or more non-ovarian cancer conditions that mayelevate serum levels of ovary related markers, including but not limitedto: cancers of the endometrium, cervix, fallopian tubes, pancreas,breast, lung and colon; nonmalignant conditions such as pregnancy,endometriosis, pelvic inflammatory disease and uterine fibroids;diagnosing conditions which have similar symptoms, signs andcomplications as ovarian cancer and where the differential diagnosisbetween them and ovarian cancer is of clinical importance including butnot limited to: non-malignant causes of pelvic mass, including, but notlimited to: benign (functional) ovarian cyst, uterine fibroids,endometriosis, benign ovarian neoplasms and inflammatory bowel lesions;determining a cause of any condition suggestive of a malignant tumorincluding but not limited to anorexia, cachexia, weight loss, fever,hypercalcemia, skeletal or abdominal pain, paraneoplastic syndrome, orascites.

In another related object the detected diseases will include autoimmuneand neoplastic disorders selected from the group consisting of Multiplesclerosis; Psoriasis; Rheumatoid arthritis; Systemic lupuserythematosus; Ulcerative colitis; Crohn's disease; immune disordersassociated with graft transplantation rejection, benign lymphocyticangiitis, lupus erythematosus, Hashimoto's thyroiditis, primarymyxedema, Graves' disease, pernicious anemia, autoimmune atrophicgastritis, Addison's disease, insulin dependent diabetes mellitis, goodpasture's syndrome, myasthenia gravis, pemphigus, sympatheticophthalmia, autoimmune uveitis, autoimmune hemolytic anemia, idiopathicthrombocytopenia, primary biliary cirrhosis, chronic action hepatitis,ulceratis colitis, Sjogren's syndrome, rheumatic disease, polymyositis,scleroderma, mixed connective tissue disease, inflammatory rheumatism,degenerative rheumatism, extra-articular rheumatism, collagen diseases,chronic polyarthritis, psoriasis arthropathica, ankylosing spondylitis,juvenile rheumatoid arthritis, periarthritis humeroscapularis,panarteriitis nodosa, progressive systemic scleroderma, arthritisuratica, dermatomyositis, muscular rheumatism, myositis, myogelosis andchondrocalcinosis.

In another related object the detected diseases will include rejectionof any organ transplant and/or Graft versus host disease.

In a related aspect the foregoing assays will detect cells affected bythe disease using the antibody that binds specifically to theAI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15),AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5(SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23), or AA424839_(—)1_P11 (SEQ IDNO:24), H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73) or R31375_P33 (SEQID NO:74) protein wherein the assays may be effected in vitro or invivo, and include RIA, ELISA, fluorimetric assays, FACS, slot blot,Western blot, immunohistochemical assays, radioimaging assays and thelike. In some embodiments, this invention provides a method fordiagnosing a disease in a subject, comprising detecting in the subjector in a sample obtained from said subject at least one polypeptide orpolynucleotide selected from the group consisting of:

a polypeptide comprising an amino acid sequence as set forth in any oneof SEQ ID NOs: 11-16, 21-34, 35-40, 43-44, 48-50, 70-76, 138-151, 296,298-302;

a polypeptide comprising a bridge, edge portion, tail or head portion,of any one of SEQ. ID NOs: 284-295, or a homologue or a fragmentthereof;

a polynucleotide comprising a nucleic acid sequence as set forth in anyone of SEQ ID NOs: 1-10, 17-20, 25-34, 41-42, 45-46, 51-69;

a polynucleotide comprising a nucleic acid sequence encoding apolypeptide comprising a bridge, edge portion, tail or head portion, ofany one of SEQ. ID NOs: 284-295;

an oligonucleotide having a nucleic acid sequence as set forth in SEQ.ID

NOs: 187, 190, 193, 196, 199, 202, 205, 208, 211, 214, 217, 220, 223,226, 229, 232, 235, 238, 241, 244, 247, 250, 253.

According to further embodiment, detecting a polypeptide of theinvention comprises employing an antibody capable of specificallybinding to at least one epitope of a polypeptide comprising an aminoacid sequence of a polypeptide comprising a bridge, edge portion, tail,or head portion of any one of SEQ. ID NOs: 284-295. According to oneembodiment, detecting the presence of the polypeptide or polynucleotideis indicative of the presence of the disease and/or its severity and/orits progress. According to another embodiment, a change in theexpression and/or the level of the polynucleotide or polypeptidecompared to its expression and/or level in a healthy subject or a sampleobtained therefrom is indicative of the presence of the disease and/orits severity and/or its progress. According to a further embodiment, achange in the expression and/or level of the polynucleotide orpolypeptide compared to its level and/or expression in said subject orin a sample obtained therefrom at earlier stage is indicative of theprogress of the disease. According to still further embodiment,detecting the presence and/or relative change in the expression and/orlevel of the polynucleotide or polypeptide is useful for selecting atreatment and/or monitoring a treatment of the disease.

According to one embodiment, detecting a polynucleotide of the inventioncomprises employing a primer pair, comprising a pair of isolatedoligonucleotides capable of specifically hybridizing to at least aportion of a polynucleotide having a nucleic acid sequence as set forthin SEQ. ID NOs: 187, 190, 193, 196, 199, 202, 205, 208, 211, 214, 217,220, 223, 226, 229, 232, 235, 238, 241, 244, 247, 250, 253, orpolynucleotides homologous thereto.

According to another embodiment, detecting a polynucleotide of theinvention comprises employing a primer pair, comprising a pair ofisolated oligonucleotides as set forth in SEQ. ID NOs:185-186, 188-189,191-192, 194-195, 197-198, 200-201, 203-204, 206-207, 209-210, 212-213,215-216, 218-219, 221-222, 224-225, 227-228, 230-231, 233-234, 236-237,239-240, 242-243, 245-246, 248-249, 251-252.

The invention also includes the following specific embodiments.

In one embodiment the invention includes an isolated polypeptideselected from AI581519_P9 (SEQ ID NO:15), AI581519_P10 (SEQ ID NO:16),AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P1 (SEQ ID NO:44),H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P14(SEQ ID NO:72), R31375_P31 (SEQ ID NO:73), R31375_P33 (SEQ ID NO:74) ora fragment or variant thereof that possesses at least 95, 96, 97, 98 or99% sequence identity therewith.

In another embodiment the invention includes a fragment or conjugatecomprising any one of the foregoing polypeptides.

In another embodiment the invention includes any one of the foregoingpolypeptides fused to an immunoglobulin domain.

In another embodiment the invention includes any of the foregoingpolypeptides attached to a detectable or therapeutic moiety.

In another embodiment the invention includes a nucleic acid sequenceencoding any of the foregoing polypeptides.

In another embodiment the invention includes any of the nucleic acidsequences selected from AI581519_T10 (SEQ ID NO:9), AI581519_T11 (SEQ IDNO:10), AA424839_(—)1_T7 (SEQ ID NO:20), H68654_(—)1_T8 (SEQ ID NO:28),H68654_(—)1_T15 (SEQ ID NO:29), H68654_(—)1_T16 (SEQ ID NO:30),H68654_(—)1_T17 (SEQ ID NO:31), H68654_(—)1_T18 (SEQ ID NO:32),H68654_(—)1_T19 (SEQ ID NO:33), H68654_(—)1_T20 (SEQ ID NO:34),AI216611_T1 (SEQ ID NO:42), H19011_(—)1_T8 (SEQ ID NO:45),H19011_(—)1_T9 (SEQ ID NO:46), R31375_T19 (SEQ ID NO:65); R31375_T25(SEQ ID NO:66), R31375_T26 (SEQ ID NO:67), R31375_T29 (SEQ ID NO:68),R31375_T39 (SEQ ID NO:69), or a fragment or variant and conjugatescontaining that possesses at least 95, 96, 97, 98 or 99% sequenceidentity therewith.

In another embodiment the invention includes an isolated VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 ectodomain polypeptide, orfragment or conjugate thereof.

In another embodiment the invention includes any of the foregoingpolypeptides, comprising a sequence of amino acid residues having atleast 95, 96, 97, 98 or 99% sequence identity with amino acid residues23-234 of AI581519_P3 (SEQ ID NO:11), corresponding to amino acidsequence depicted in SEQ ID NO:138, or amino acid residues 23-270 ofAI581519_P4 (SEQ ID NO:12), corresponding to amino acid sequencedepicted in SEQ ID NO:139, or amino acid residues 23-296 of AI581519_P5(SEQ ID NO:13), corresponding to amino acid sequence depicted in SEQ IDNO:140, or amino acid residues 23-193 of AI581519_P7 (SEQ ID NO:14),corresponding to amino acid sequence depicted in SEQ ID NO:141, or aminoacid residues 23-203 of AI581519_P9 (SEQ ID NO:15), corresponding toamino acid sequence depicted in SEQ ID NO:142, or amino acid residues23-231 of AI581519_P10 (SEQ ID NO:16), corresponding to amino acidsequence depicted in SEQ ID NO:143, or residues 26-293 of AI581519_P5(SEQ ID NO:13), corresponding to amino acid sequence depicted in SEQ IDNO:302, or amino acid residues 24-162 of AA424839_P3 (SEQ ID NO:22), orAA424839_P5 (SEQ ID NO:21), corresponding to amino acid sequencedepicted in SEQ ID NO:75, or amino acid residues 24-456 of AA424839_P7(SEQ ID NO:23), corresponding to amino acid sequence depicted in SEQ IDNO:76, or amino acid residues 24-105 of AA424839_(—)1_P11 (SEQ IDNO:24), corresponding to amino acid sequence depicted in SEQ ID NO:296,or residues 50-160 of AA424839_(—)1_P3 (SEQ ID NO:22), corresponding toamino acid sequence depicted in SEQ ID NO:301, or amino acid residues38-349 of H68654_(—)1_P2 (SEQ ID NO:35), corresponding to amino acidsequence depicted in SEQ ID NO:144, or residues 19-337 of H68654_(—)1_P5(SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ IDNO:38), H68654_(—)1_P13 (SEQ ID NO:39), or H68654_(—)1_P14 (SEQ IDNO:40), corresponding to amino acid sequence depicted in SEQ ID NO:145,or residues 1-335 of the sequences H68654_(—)1_P5 (SEQ ID NO:36),corresponding to amino acid sequence depicted in SEQ ID NO:300, or aminoacid residues 29-147 of the sequence AI216611_P0 (SEQ ID NO:43) orAI216611_P1 (SEQ ID NO:44), or residues 1-145 of the sequenceAI216611_P0 (SEQ ID NO:43), corresponding to amino acid sequencedepicted in SEQ ID NO:298, corresponding to amino acid sequence depictedin SEQ ID NO:146, or amino acid residues 21-186 of H19011_(—)1_P8 (SEQID NO:48), corresponding to amino acid sequence depicted in SEQ IDNO:147, or residues 21-169 of H19011_(—)1_P9 (SEQ ID NO:50),corresponding to amino acid sequence depicted in SEQ ID NO:148, orresidues 1-184 of the sequence H19011_(—)1_P8 (SEQ ID NO:48),corresponding to amino acid sequence depicted in SEQ ID NO:299, or aminoacid residues 21-36 of R31375_P0 (SEQ ID NO:70) or R31375_P31 (SEQ IDNO:73), corresponding to amino acid sequence depicted in SEQ ID NO:149,or residues 21-65 of R31375_P14 (SEQ ID NO:72), corresponding to aminoacid sequence depicted in SEQ ID NO:150, or residues 21-25 of R31375_P33(SEQ ID NO:74), corresponding to amino acid sequence depicted in SEQ IDNO:151, or residues 1-63 of the sequence R31375_P14 (SEQ ID NO:72),corresponding to amino acid sequence depicted in SEQ ID NO:297.

In another embodiment the invention includes any of the foregoingpolypeptides, comprising the extracellular domain of AI581519_P3 (SEQ IDNO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13),AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15), AI581519_P10(SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21),AA424839_P7 (SEQ ID NO:23), AA424839_(—)1_P11 (SEQ ID NO:24),H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73) or R31375_P33 (SEQID NO:74).

In another embodiment the invention includes any of the foregoingpolypeptides, attached to a detectable or therapeutic moiety.

In another embodiment the invention includes any of the foregoingnucleic acid sequences encoding any one of the VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, FXYD3 ectodomain polypeptides and conjugatescontaining.

In another embodiment the invention includes an expression vectorcontaining any of the foregoing nucleic acid sequences.

In another embodiment the invention includes a host cell comprising theforegoing expression vector or a virus containing a nucleic acidsequence encoding the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3ectodomain polypeptide, or fragment or conjugate thereof, wherein thecell expresses the polypeptide encoded by the DNA segment.

In another embodiment the invention includes a method of producinganyone of the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3ectodomain polypeptides, or fragment or conjugate thereof, comprisingculturing the foregoing host cell, wherein the cell expresses thepolypeptide encoded by the DNA segment or nucleic acid and recoveringsaid polypeptide.

In another embodiment the invention includes any of the foregoingisolated soluble VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3ectodomain wherein said polypeptide blocks or inhibits the interactionof AI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5(SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15),AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5(SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23), AA424839_(—)1_P11 (SEQ IDNO:24), H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73), R31375_P33 (SEQ IDNO:74), or a fragment or variant thereof with a corresponding functionalcounterpart.

In another embodiment the invention includes the foregoing isolatedsoluble VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 ectodomains,wherein said polypeptide replaces or augments the interaction ofAI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15),AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5(SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23), AA424839_(—)1_P11 (SEQ IDNO:24), H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73) or R31375_P33 (SEQID NO:74), or a fragment or variant or conjugate thereof with acorresponding functional counterpart.

In another embodiment the invention includes a fusion protein comprisingany of the foregoing isolated soluble VSIG1, ILDR1, LOC253012, AI216611,C1ORF32, FXYD3 ectodomain joined to a non-VSIG1, non-ILDR1,non-LOC253012, non-AI216611, non-C1ORF32, non-FXYD3 protein sequence,correspondingly.

In another embodiment the invention includes any of the foregoing fusionproteins, wherein the non-VSIG1, non-ILDR1, non-LOC253012, non-AI216611,non-C1ORF32, non-FXYD3 protein is at least a portion of animmunoglobulin molecule.

In another embodiment the invention includes any of the foregoing fusionproteins, wherein a polyalkyl oxide moiety such as polyethylene glycolis attached to the polypeptide.

In another embodiment the invention includes any of the foregoing fusionproteins, wherein the immunoglobulin heavy chain constant region is anFc fragment.

In another embodiment the invention includes any one of the proteinsequences of the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 and FXYD3ECDs fused to mouse Fc, as set forth in any one of amino acid sequencesas depicted in SEQ ID NOs: 103-108, or nucleic acid sequences encodingthe VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 and FXYD3 ECDs fused tomouse Fc. The invention further includes the nucleic acid sequencesencoding the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 and FXYD3 ECDsfused to mouse Fc, as set forth in any one of nucleic acid sequencesdepicted in SEQ ID NOs:97-102.

In another embodiment the invention includes any of the foregoing fusionproteins wherein the immunoglobulin heavy chain constant region is anisotype selected from the group consisting of an IgG1, IgG2, IgG3, IgG4,IgM, IgE, IgA and IgD.

In another embodiment the invention includes any of the foregoing fusionproteins, wherein the polypeptide is fused to a VASP domain.

In another embodiment the invention includes any of the foregoing fusionproteins, wherein the fusion protein modulates lymphocyte activation.

In another embodiment the invention includes a pharmaceuticalcomposition comprising any of the foregoing polynucleotide sequences andfurther comprising a pharmaceutically acceptable diluent or carrier.

In another embodiment the invention includes a pharmaceuticalcomposition comprising the foregoing vector and further comprising apharmaceutically acceptable diluent or carrier.

In another embodiment the invention includes a pharmaceuticalcomposition comprising the foregoing host cell and further comprising apharmaceutically acceptable diluent or carrier.

In another embodiment the invention includes a pharmaceuticalcomposition comprising any of the foregoing VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, FXYD3 ectodomains and further comprising apharmaceutically acceptable diluent or carrier.

In another embodiment the invention includes a pharmaceuticalcomposition comprising any of the foregoing polypeptides and furthercomprising a pharmaceutically acceptable diluent or carrier.

In another embodiment the invention includes a pharmaceuticalcomposition comprising the foregoing fusion protein and furthercomprising a pharmaceutically acceptable diluent or carrier.

In another embodiment the invention includes a method for treating orpreventing cancer, comprising administering to a subject in need thereofa pharmaceutical composition comprising: a soluble molecule having theextracellular domain of VSIG1, ILDR1, LOC253012, AI216611, C1ORF32,FXYD3 polypeptide, or fragment or conjugate thereof; or polypeptide,comprising a sequence of amino acid residues having at least 95, 96, 97,98 or 99% sequence identity with amino acid residues 23-234 ofAI581519_P3 (SEQ ID NO:11), corresponding to amino acid sequencedepicted in SEQ ID NO:138, or amino acid residues 23-270 of AI581519_P4(SEQ ID NO:12), corresponding to amino acid sequence depicted in SEQ IDNO:139, or amino acid residues 23-296 of AI581519_P5 (SEQ ID NO:13),corresponding to amino acid sequence depicted in SEQ ID NO:140, or aminoacid residues 23-193 of AI581519_P7 (SEQ ID NO:14), corresponding toamino acid sequence depicted in SEQ ID NO:141, or amino acid residues23-203 of AI581519_P9 (SEQ ID NO:15), corresponding to amino acidsequence depicted in SEQ ID NO:142, or amino acid residues 23-231 ofAI581519_P10 (SEQ ID NO:16), corresponding to amino acid sequencedepicted in SEQ ID NO:143, or residues 26-293 of AI581519_P5 (SEQ IDNO:13), corresponding to amino acid sequence depicted in SEQ ID NO:302,or amino acid residues 24-162 of AA424839_P3 (SEQ ID NO:22), orAA424839_P5 (SEQ ID NO:21), corresponding to amino acid sequencedepicted in SEQ ID NO:75, or amino acid residues 24-456 of AA424839_P7(SEQ ID NO:23), corresponding to amino acid sequence depicted in SEQ IDNO:76, or amino acid residues 24-105 of AA424839_(—)1_P11 (SEQ IDNO:24), corresponding to amino acid sequence depicted in SEQ ID NO:296,or residues 50-160 of AA424839_(—)1_P3 (SEQ ID NO:22), corresponding toamino acid sequence depicted in SEQ ID NO:301, or amino acid residues38-349 of H68654_(—)1_P2 (SEQ ID NO:35), corresponding to amino acidsequence depicted in SEQ ID NO:144, or residues 19-337 of H68654_(—)1_P5(SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ IDNO:38), H68654_(—)1_P13 (SEQ ID NO:39), or H68654_(—)1_P14 (SEQ IDNO:40), corresponding to amino acid sequence depicted in SEQ ID NO:145,or residues 1-335 of the sequences H68654_(—)1_P5 (SEQ ID NO:36),corresponding to amino acid sequence depicted in SEQ ID NO:300, or aminoacid residues 29-147 of the sequence AI216611_P0 (SEQ ID NO:43) orAI216611_P1 (SEQ ID NO:44), or residues 1-145 of the sequenceAI216611_P0 (SEQ ID NO:43), corresponding to amino acid sequencedepicted in SEQ ID NO:298, corresponding to amino acid sequence depictedin SEQ ID NO:146, or amino acid residues 21-186 of H19011_(—)1_P8 (SEQID NO:48), corresponding to amino acid sequence depicted in SEQ IDNO:147, or residues 21-169 of H19011_(—)1_P9 (SEQ ID NO:50),corresponding to amino acid sequence depicted in SEQ ID NO:148, orresidues 1-184 of the sequence H19011_(—)1_P8 (SEQ ID NO:48),corresponding to amino acid sequence depicted in SEQ ID NO:299, or aminoacid residues 21-36 of R31375_P0 (SEQ ID NO:70) or R31375_P31 (SEQ IDNO:73), corresponding to amino acid sequence depicted in SEQ ID NO:149,or residues 21-65 of R31375_P14 (SEQ ID NO:72), corresponding to aminoacid sequence depicted in SEQ ID NO:150, or residues 21-25 of R31375_P33(SEQ ID NO:74), corresponding to amino acid sequence depicted in SEQ IDNO:151, or residues 1-63 of the sequence R31375_P14 (SEQ ID NO:72),corresponding to amino acid sequence depicted in SEQ ID NO:297, or anucleic acid sequence encoding the same.

In another embodiment the invention includes the foregoing method,wherein the cancer is selected from a group consisting of hematologicalmalignancies such as acute lymphocytic leukemia, chronic lymphocyticleukemia, acute myelogenous leukemia, chronic myelogenous leukemia,multiple myeloma, Hodgkin's lymphoma, Non-Hodgkin's lymphoma, and softor solid tumors such as cancer of breast, prostate, lung, ovary, colon,spleen, kidney, bladder, head and neck, uterus, testicles, stomach,cervix, liver, bone, skin, pancreas, brain and wherein the cancer isnon-metastatic, invasive or metastatic.

In another embodiment the invention includes the foregoing methodwherein the cancer is selected from the group consisting of lung cancer,ovarian cancer or colon cancer, and wherein the lung cancer, the ovariancancer or the colon cancer is non-metastatic, invasive or metastatic.

In another embodiment the invention includes a method for treating orpreventing immune related conditions, such as autoimmune diseases ortransplant rejection, comprising administering to a subject in needthereof a pharmaceutical composition comprising: a soluble moleculehaving the extracellular domain of VSIG1, ILDR1, LOC253012, AI216611,C1ORF32, FXYD3 polypeptide, or fragment or conjugate thereof; orpolypeptide, comprising a sequence of amino acid residues having atleast 95, 96, 97, 98 or 99% sequence identity with amino acid residues23-234 of AI581519_P3 (SEQ ID NO:11), corresponding to amino acidsequence depicted in SEQ ID NO:138, or amino acid residues 23-270 ofAI581519_P4 (SEQ ID NO:12), corresponding to amino acid sequencedepicted in SEQ ID NO:139, or amino acid residues 23-296 of AI581519_P5(SEQ ID NO:13), corresponding to amino acid sequence depicted in SEQ IDNO:140, or amino acid residues 23-193 of AI581519_P7 (SEQ ID NO:14),corresponding to amino acid sequence depicted in SEQ ID NO:141, or aminoacid residues 23-203 of AI581519_P9 (SEQ ID NO:15), corresponding toamino acid sequence depicted in SEQ ID NO:142, or amino acid residues23-231 of AI581519_P10 (SEQ ID NO:16), corresponding to amino acidsequence depicted in SEQ ID NO:143, or residues 26-293 of AI581519_P5(SEQ ID NO:13), corresponding to amino acid sequence depicted in SEQ IDNO:302, or amino acid residues 24-162 of AA424839_P3 (SEQ ID NO:22), orAA424839_P5 (SEQ ID NO:21), corresponding to amino acid sequencedepicted in SEQ ID NO:75, or amino acid residues 24-456 of AA424839_P7(SEQ ID NO:23), corresponding to amino acid sequence depicted in SEQ IDNO:76, or amino acid residues 24-105 of AA424839_(—)1_P11 (SEQ IDNO:24), corresponding to amino acid sequence depicted in SEQ ID NO:296,or residues 50-160 of AA424839_(—)1_P3 (SEQ ID NO:22), corresponding toamino acid sequence depicted in SEQ ID NO:301, or amino acid residues38-349 of H68654_(—)1_P2 (SEQ ID NO:35), corresponding to amino acidsequence depicted in SEQ ID NO:144, or residues 19-337 of H68654_(—)1_P5(SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ IDNO:38), H68654_(—)1_P13 (SEQ ID NO:39), or H68654_(—)1_P14 (SEQ IDNO:40), corresponding to amino acid sequence depicted in SEQ ID NO:145,or residues 1-335 of the sequences H68654_(—)1_P5 (SEQ ID NO:36),corresponding to amino acid sequence depicted in SEQ ID NO:300, or aminoacid residues 29-147 of the sequence AI216611_P0 (SEQ ID NO:43) orAI216611_P1 (SEQ ID NO:44), or residues 1-145 of the sequenceAI216611_P0 (SEQ ID NO:43), corresponding to amino acid sequencedepicted in SEQ ID NO:298, corresponding to amino acid sequence depictedin SEQ ID NO:146, or amino acid residues 21-186 of H19011_(—)1_P8 (SEQID NO:48), corresponding to amino acid sequence depicted in SEQ IDNO:147, or residues 21-169 of H19011_(—)1_P9 (SEQ ID NO:50),corresponding to amino acid sequence depicted in SEQ ID NO:148, residues1-184 of the sequence H19011_(—)1_P8 (SEQ ID NO:48), corresponding toamino acid sequence depicted in SEQ ID NO:299, or residues 21-36 ofR31375_P0 (SEQ ID NO:70) or R31375_P31 (SEQ ID NO:73), corresponding toamino acid sequence depicted in SEQ ID NO:149, or residues 21-65 ofR31375_P14 (SEQ ID NO:72), corresponding to amino acid sequence depictedin SEQ ID NO:150, or residues 21-25 of R31375_P33 (SEQ ID NO:74),corresponding to amino acid sequence depicted in SEQ ID NO:151, orresidues 1-63 of R31375_P14 (SEQ ID NO:72), corresponding to amino acidsequence depicted in SEQ ID NO:297, or a nucleic acid sequence encodingthe same.

In another embodiment the invention includes the foregoing method,wherein the autoimmune diseases are selected from a group consisting ofmultiple sclerosis; psoriasis; rheumatoid arthritis; systemic lupuserythematosus; ulcerative colitis; Crohn's disease; immune disordersassociated with graft transplantation rejection; benign lymphocyticangiitis, lupus erythematosus, Hashimoto's thyroiditis, primarymyxedema, Graves' disease, pernicious anemia, autoimmune atrophicgastritis, Addison's disease, insulin dependent diabetes mellitis, Goodpasture's syndrome, myasthenia gravis, pemphigus, sympatheticophthalmia, autoimmune uveitis, autoimmune hemolytic anemia, idiopathicthrombocytopenia, primary biliary cirrhosis, chronic action hepatitis,ulceratis colitis, Sjogren's syndrome, rheumatic disease, polymyositis,scleroderma, mixed connective tissue disease, inflammatory rheumatism,degenerative rheumatism, extra-articular rheumatism, collagen diseases,chronic polyarthritis, psoriasis arthropathica, ankylosing spondylitis,juvenile rheumatoid arthritis, periarthritis humeroscapularis,panarteriitis nodosa, progressive systemic scleroderma, arthritisuratica, dermatomyositis, muscular rheumatism, myositis, myogelosis andchondrocalcinosis.

In another embodiment the invention includes the foregoing method,wherein the immune related disorders are selected from transplantrejection or graft versus host disease.

In another embodiment the invention includes an siRNA, antisense RNA, orribozyme that binds the transcript encoding any one of the VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, FXYD3 polypeptides, selected fromAI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15),AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5(SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23), AA424839_(—)1_P11 (SEQ IDNO:24), H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73), R31375_P33 (SEQ IDNO:74), or a fragment or a variant thereof, and inhibits its expression.

In another embodiment the invention includes a polyclonal or monoclonalantibody that specifically binds and/or modulates an activity elicitedby any one of the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3polypeptides, selected from AI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQID NO:12), AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14),AI581519_P9 (SEQ ID NO:15), AI581519_P10 (SEQ ID NO:16), AA424839_P3(SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23),AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ ID NO:35),H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73), R31375_P33 (SEQ ID NO:74), or a fragment or a variantthereof and conjugates containing.

In another embodiment the invention includes a monoclonal or polyclonalantibody or an antigen binding fragment thereof comprising an antigenbinding site that binds specifically to any one of the VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, FXYD3 polypeptides comprised inAI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15),AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5(SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23), AA424839_(—)1_P11 (SEQ IDNO:24), H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73), R31375_P33 (SEQ IDNO:74), or fragment or variant thereof that is at least 80% identicalthereto.

In another embodiment the invention includes any of the foregoingantibodies or fragments thereof, wherein said antibody blocks orinhibits the interaction of one of AI581519_P3 (SEQ ID NO:11),AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQID NO:14), AI581519_P9 (SEQ ID NO:15), AI581519_P10 (SEQ ID NO:16),AA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQID NO:23), AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ IDNO:35), H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73), R31375_P33 (SEQ ID NO:74), or a fragment or variantthereof with a counterpart activity or function.

In another embodiment the invention includes any of the foregoingantibodies or fragments wherein said antibody replaces or augments theinteraction of AI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12),AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQID NO:15), AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22),AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23),AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ ID NO:35),H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73), R31375_P33 (SEQ ID NO:74), or a fragment or variantthereof with a counterpart function or activity.

In another embodiment the invention includes a method for modulatinglymphocyte activity, comprising contacting a AI581519_P3 (SEQ ID NO:11),AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQID NO:14), AI581519_P9 (SEQ ID NO:15), AI581519_P10 (SEQ ID NO:16),AA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQID NO:23), AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ IDNO:35), H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73), R31375_P33 (SEQ ID NO:74) positive lymphocyte with abioactive agent capable of modulating VSIG1-mediated, ILDR1-mediated,LOC253012-mediated, AI216611-mediated, C1ORF32-mediated, orFXYD3-mediated signaling in an amount effective to modulate at least onelymphocyte activity.

In another embodiment the invention includes the foregoing method,wherein said agent comprises an antagonist of VSIG1-mediated,ILDR1-mediated, LOC253012-mediated, AI216611-mediated, C1ORF32-mediatedsignaling, or FXYD3-mediated signaling and wherein said contactinginhibits the attenuation of lymphocyte activity mediated by suchsignaling.

In another embodiment the invention includes the foregoing method,wherein said contacting increases lymphocyte activity.

In another embodiment the invention includes the foregoing methodwherein said antagonist comprises a blocking agent capable ofinterfering with the functional interaction of VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, FXYD3 antigen and its counterpart.

In another embodiment the invention includes the foregoing antibody orfragment which is suitable for treatment or prevention of cancer bymodulating the activity of any one of the VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, FXYD3 proteins in a B7-like co-stimulatory system.

In another embodiment the invention includes the foregoing methodwherein the administered antibody or fragment inhibits negativestimulation of T cell activity against cancer cells.

In another embodiment the invention includes any of the foregoingantibodies or fragments, wherein the cancer is selected from the groupconsisting of hematological malignancies such as acute lymphocyticleukemia, chronic lymphocytic leukemia, acute myelogenous leukemia,chronic myelogenous leukemia, multiple myeloma, Hodgkin's lymphoma,Non-Hodgkin's lymphoma, and soft tissue or solid tumors such as cancerof breast, prostate, lung, ovary, colon, spleen, kidney, bladder, headand neck, uterus, testicles, stomach, cervix, liver, bone, skin,pancreas, brain and wherein the cancer is non-metastatic, invasive ormetastatic.

In another embodiment the invention includes any of the foregoingantibodies or fragments, which are suitable for treatment or preventionof immune related disorders, such as autoimmune diseases or transplantrejection, by modulating the activity of anyone of the VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, FXYD3 proteins in a B7-like co-stimulatorysystem.

In another embodiment the invention includes any of the foregoingantibodies or fragments, which are suitable for treating an autoimmunedisease selected from multiple sclerosis; psoriasis; rheumatoidarthritis; Systemic lupus erythematosus; ulcerative colitis; Crohn'sdisease, immune disorders associated with graft transplantationrejection, benign lymphocytic angiitis, lupus erythematosus, Hashimoto'sthyroiditis, primary myxedema, Graves' disease, pernicious anemia,autoimmune atrophic gastritis, Addison's disease, insulin dependentdiabetes mellitus, Good pasture's syndrome, myasthenia gravis,pemphigus, sympathetic ophthalmia, autoimmune uveitis, autoimmunehemolytic anemia, idiopathic thrombocytopenia, primary biliarycirrhosis, chronic action hepatitis, Sjogren's syndrome, rheumaticdisease, polymyositis, scleroderma, mixed connective tissue disease,inflammatory rheumatism, degenerative rheumatism, extra-articularrheumatism, collagen diseases, chronic polyarthritis, psoriasisarthropathica, ankylosing spondylitis, juvenile rheumatoid arthritis,periarthritis humeroscapularis, panarteriitis nodosa, progressivesystemic scleroderma, arthritis uratica, dermatomyositis, muscularrheumatism, myositis, myogelosis and chondrocalcinosis.

In another embodiment the invention includes any of the foregoingantibodies or fragments, suitable for treating transplant rejection orgraft versus host disease.

In another embodiment the invention includes any of the foregoingantibodies or fragments, that specifically binds to amino-acids: 23-234of AI581519_P3 (SEQ ID NO:11), corresponding to amino acid sequencedepicted in SEQ ID NO:138, or amino acid residues 23-270 of AI581519_P4(SEQ ID NO:12), corresponding to amino acid sequence depicted in SEQ IDNO:139, or amino acid residues 23-296 of AI581519_P5 (SEQ ID NO:13),corresponding to amino acid sequence depicted in SEQ ID NO:140, or aminoacid residues 23-193 of AI581519_P7 (SEQ ID NO:14), corresponding toamino acid sequence depicted in SEQ ID NO:141, or amino acid residues23-203 of AI581519_P9 (SEQ ID NO:15), corresponding to amino acidsequence depicted in SEQ ID NO:142, or amino acid residues 23-231 ofAI581519_P10 (SEQ ID NO:16), corresponding to amino acid sequencedepicted in SEQ ID NO:143, or residues 26-293 of AI581519_P5 (SEQ IDNO:13), corresponding to amino acid sequence depicted in SEQ ID NO:302,or amino acid residues 24-162 of AA424839_P3 (SEQ ID NO:22), orAA424839_P5 (SEQ ID NO:21), corresponding to amino acid sequencedepicted in SEQ ID NO:75, or amino acid residues 24-456 of AA424839_P7(SEQ ID NO:23), corresponding to amino acid sequence depicted in SEQ IDNO:76, or amino acid residues 24-105 of AA424839_(—)1_P11 (SEQ IDNO:24), corresponding to amino acid sequence depicted in SEQ ID NO:296,or residues 50-160 of AA424839_(—)1_P3 (SEQ ID NO:22), corresponding toamino acid sequence depicted in SEQ ID NO:301, or amino acid residues38-349 of H68654_(—)1_P2 (SEQ ID NO:35), corresponding to amino acidsequence depicted in SEQ ID NO:144, or residues 19-337 of H68654_(—)1_P5(SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ IDNO:38), H68654_(—)1_P13 (SEQ ID NO:39), or H68654_(—)1_P14 (SEQ IDNO:40), corresponding to amino acid sequence depicted in SEQ ID NO:145,or residues 1-335 of the sequences H68654_(—)1_P5 (SEQ ID NO:36),corresponding to amino acid sequence depicted in SEQ ID NO:300, or aminoacid residues 29-147 of the sequence AI216611_P0 (SEQ ID NO:43) orAI216611_P1 (SEQ ID NO:44), or residues 1-145 of the sequenceAI216611_P0 (SEQ ID NO:43), corresponding to amino acid sequencedepicted in SEQ ID NO:298, corresponding to amino acid sequence depictedin SEQ ID NO:146, or amino acid residues 21-186 of H19011_(—)1_P8 (SEQID NO:48), corresponding to amino acid sequence depicted in SEQ IDNO:147, or residues 21-169 of H19011_(—)1_P9 (SEQ ID NO:50),corresponding to amino acid sequence depicted in SEQ ID NO:148, orresidues 1-184 of the sequence H19011_(—)1_P8 (SEQ ID NO:48),corresponding to amino acid sequence depicted in SEQ ID NO:299, or aminoacid residues 21-36 of R31375_P0 (SEQ ID NO:70) or R31375_P31 (SEQ IDNO:73), corresponding to amino acid sequence depicted in SEQ ID NO:149,or residues 21-65 of R31375_P14 (SEQ ID NO:72), corresponding to aminoacid sequence depicted in SEQ ID NO:150, or residues 21-25 of R31375_P33(SEQ ID NO:74), corresponding to amino acid sequence depicted in SEQ IDNO:151, or residues 1-63 of the sequence R31375_P14 (SEQ ID NO:72),corresponding to amino acid sequence depicted in SEQ ID NO:297, or avariant or fragment or an epitope thereof.

In another embodiment the invention includes any of the foregoingantibodies or fragments, wherein the antigen binding site contains fromabout 3-7 contiguous or non-contiguous amino acids, more typically atleast 5 contiguous or non-contiguous amino acids. These binding sitesinclude conformational and non-conformational epitopes.

In another embodiment the invention includes any of the foregoingantibodies or fragments, wherein the antibody is a fully human antibody.

In another embodiment the invention includes any of the foregoingantibodies or fragments, wherein the antibody is a chimeric antibody.

In another embodiment the invention includes the foregoing antibodies orfragments wherein the antibody is a humanized or primatized antibody.

In another embodiment the invention includes any of the foregoingantibodies or fragments, wherein the fragment is selected from the groupconsisting of Fab, Fab′, F(ab′)2, F(ab′), F(ab), Fv or scFv fragment andminimal recognition unit.

In another embodiment the invention includes any of the foregoingantibodies or fragments, wherein the antibody or fragment is coupled toa detectable marker, or to an effector moiety.

In another embodiment the invention includes any of the foregoingantibodies or fragments, wherein the effector moiety is an enzyme, atoxin, a therapeutic agent, or a chemotherapeutic agent.

In another embodiment the invention includes any of the foregoingantibodies or fragments, wherein the detectable marker is aradioisotope, a metal chelator, an enzyme, a fluorescent compound, abioluminescent compound or a chemiluminescent compound.

In another embodiment the invention includes a pharmaceuticalcomposition that comprises any of the foregoing antibodies or a fragmentthereof.

In another embodiment the invention includes a pharmaceuticalcomposition that comprises the foregoing antibodies or a fragmentthereof.

In another embodiment the invention includes a method of inducing orenhancing an immune response, comprising administering to a patient inneed thereof any of the foregoing antibodies or fragments and detectinginduction or enhancement of said immune response.

In another embodiment the invention includes a method for potentiating asecondary immune response to an antigen in a patient, which methodcomprises administering effective amounts any of the foregoingantibodies or fragments.

In another embodiment the invention includes the foregoing method,wherein the antigen is preferably a cancer antigen, a viral antigen or abacterial antigen, and the patient has preferably received treatmentwith an anticancer vaccine or a viral vaccine.

In another embodiment the invention includes a method of treating apatient with a VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3positive malignancy, comprising administering to the patient aneffective amount of any of the foregoing antibodies or fragments.

In another embodiment the invention includes the foregoing methodfurther comprising co-administering a chemotherapeutic agent.

In another embodiment the invention includes the foregoing method,wherein said malignancy is selected from a group consisting ofhematological malignancies such as acute lymphocytic leukemia, chroniclymphocytic leukemia, acute myelogenous leukemia, chronic myelogenousleukemia, multiple myeloma, Hodgkin's lymphoma, Non-Hodgkin's lymphoma,and soft or solid tumors such as cancer of breast, prostate, lung,ovary, colon, spleen, kidney, bladder, head and neck, uterus, testicles,stomach, cervix, liver, bone, skin, pancreas, brain and wherein thecancer is non-metastatic, invasive or metastatic.

In another embodiment the invention includes the foregoing method,wherein said malignancy is selected from the group consisting of lungcancer, ovarian cancer, colon cancer, and wherein the lung cancer, theovarian cancer or the colon cancer is non-metastatic, invasive ormetastatic.

In another embodiment the invention includes an assay for detecting thepresence of AI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12),AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQID NO:15), AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22),AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23),AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ ID NO:35),H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73), R31375_P33 (SEQ ID NO:74), or a fragment or variantthereof in a biological sample comprising contacting the sample with anantibody of any one of the foregoing, and detecting the binding ofAI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15),AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5(SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23), AA424839_(—)1_P11 (SEQ IDNO:24), H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73), R31375_P33 (SEQ IDNO:74), or a fragment or variant thereof in the sample.

In another embodiment the invention includes a method for detecting adisease, diagnosing a disease, monitoring disease progression ortreatment efficacy or relapse of a disease, or selecting a therapy for adisease, comprising detecting expression of a AI581519_P3 (SEQ IDNO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13),AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15), AI581519_P10(SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21),AA424839_P7 (SEQ ID NO:23), AA424839_(—)1_P11 (SEQ ID NO:24),H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73), R31375_P33 (SEQ IDNO:74), or a fragment or variant thereof.

In another embodiment the invention includes the foregoing methodwherein detecting expression AI581519_P3 (SEQ ID NO:11), AI581519_P4(SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14),AI581519_P9 (SEQ ID NO:15), AI581519_P10 (SEQ ID NO:16), AA424839_P3(SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23),AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ ID NO:35),H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73), R31375_P33 (SEQ ID NO:74), or a fragment or variantthereof is performed in vivo or in vitro.

In another embodiment the invention includes the foregoing method,wherein the disease is selected from lung cancer, ovarian cancer, orcolon cancer, and wherein the lung cancer, the ovarian cancer or thecolon cancer is non-metastatic, invasive or metastatic.

In another embodiment the invention includes the foregoing method,wherein the disease is multiple sclerosis; psoriasis; rheumatoidarthritis; Systemic lupus erythematosus; ulcerative colitis; Crohn'sdisease; immune disorders associated with graft transplantationrejection, benign lymphocytic angiitis, lupus erythematosus, Hashimoto'sthyroiditis, primary myxedema, Graves' disease, pernicious anemia,autoimmune atrophic gastritis, Addison's disease, insulin dependentdiabetes mellitis, good pasture's syndrome, myasthenia gravis,pemphigus, sympathetic ophthalmia, autoimmune uveitis, autoimmunehemolytic anemia, idiopathic thrombocytopenia, primary biliarycirrhosis, chronic action hepatitis, ulceratis colitis, Sjogren'ssyndrome, rheumatic disease, polymyositis, scleroderma, mixed connectivetissue disease, inflammatory rheumatism, degenerative rheumatism,extra-articular rheumatism, collagen diseases, chronic polyarthritis,psoriasis arthropathica, ankylosing spondylitis, juvenile rheumatoidarthritis, periarthritis humeroscapularis, panarteriitis nodosa,progressive systemic scleroderma, arthritis uratica, dermatomyositis,muscular rheumatism, myositis, myogelosis or chondrocalcinosis.

In another embodiment the invention includes a method of inhibitinggrowth of cells that express a polypeptide selected from AI581519_P3(SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13),AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15), AI581519_P10(SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21),AA424839_P7 (SEQ ID NO:23), AA424839_(—)1_P11 (SEQ ID NO:24),H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73), R31375_P33 (SEQ IDNO:74), or a fragment or variant thereof in a subject, comprising:administering to said subject any of the foregoing antibodies orfragments.

In another embodiment the invention includes a method of treating orpreventing cancer comprising the administration of a therapeuticallyeffective amount of an antibody or binding fragment that specificallybinds the AI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12),AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQID NO:15), AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22),AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23),AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ ID NO:35),H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73), R31375_P33 (SEQ ID NO:74), or a fragment or variantthereof that possesses at least 80% sequence identity therewith.

In another embodiment the invention includes the foregoing method,wherein the cancer is selected from a group consisting of hematologicalmalignancies such as acute lymphocytic leukemia, chronic lymphocyticleukemia, acute myelogenous leukemia, chronic myelogenous leukemia,multiple myeloma, Hodgkin's lymphoma, Non-Hodgkin's lymphoma, and softor solid tumors such as cancer of breast, prostate, lung, ovary, colon,spleen, kidney, bladder, head and neck, uterus, testicles, stomach,cervix, liver, bone, skin, pancreas, brain and wherein the cancer isnon-metastatic, invasive or metastatic.

In another embodiment the invention includes the foregoing method,wherein the cancer is selected from the group consisting of lung cancer,ovarian cancer, or colon cancer, and wherein the lung cancer, theovarian cancer or the colon cancer is non-metastatic, invasive ormetastatic.

In another embodiment the invention includes the foregoing methodwherein the antibody is a human, humanized or chimeric antibody orantigen binding fragment.

In another embodiment the invention includes the foregoing methodwherein the antibody or fragment is attached directly or indirectly toan effector moiety.

In another embodiment the invention includes the foregoing method,wherein the effector is selected from a drug, toxin, radionuclide,fluorophore and an enzyme.

In another embodiment the invention includes a method for treating orpreventing an immune disorder, such as autoimmune or transplant relateddisease, comprising administering to a patient a therapeuticallyeffective amount of an antibody that specifically binds to AI581519_P3(SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13),AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15), AI581519_P10(SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21),AA424839_P7 (SEQ ID NO:23), AA424839_(—)1_P11 (SEQ ID NO:24),H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73), R31375_P33 (SEQ IDNO:74),

or a fragment or variant thereof that possesses at least 80% sequenceidentity therewith.

In another embodiment the invention includes the foregoing method,wherein the antibody has an antigen-binding region specific for theextracellular domain of any one of said VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, FXYD3 polypeptides.

In another embodiment the invention includes the foregoing method,wherein the antibody or fragment modulates the B7/co-stimulatory systemin a manner that inhibits positive stimulation of T cell activity thatcreated an autoimmune effect.

In another embodiment the invention includes the foregoing method,wherein the treatment is combined with a moiety useful for treatingautoimmune or transplant rejection conditions.

In another embodiment the invention includes the foregoing method,wherein the moiety is a cytokine antibody, cytokine receptor antibody,drug, or another immunomodulatory agent.

In another embodiment the invention includes the foregoing method,wherein the autoimmune diseases are selected from a group consisting ofmultiple sclerosis; psoriasis; rheumatoid arthritis; systemic lupuserythematosus; ulcerative colitis; Crohn's disease; immune disordersassociated with graft transplantation rejection, benign lymphocyticangiitis, lupus erythematosus, Hashimoto's thyroiditis, primarymyxedema, Graves' disease, pernicious anemia, autoimmune atrophicgastritis, Addison's disease, insulin dependent diabetes mellitis, Goodpasture's syndrome, myasthenia gravis, pemphigus, sympatheticophthalmia, autoimmune uveitis, autoimmune hemolytic anemia, idiopathicthrombocytopenia, primary biliary cirrhosis, chronic action hepatitis,ulceratis colitis, Sjogren's syndrome, rheumatic disease, polymyositis,scleroderma, mixed connective tissue disease, inflammatory rheumatism,degenerative rheumatism, extra-articular rheumatism, collagen diseases,chronic polyarthritis, psoriasis arthropathica, ankylosing spondylitis,juvenile rheumatoid arthritis, periarthritis humeroscapularis,panarteriitis nodosa, progressive systemic scleroderma, arthritisuratica, dermatomyositis, muscular rheumatism, myositis, myogelosis andchondrocalcinosis.

In another embodiment the invention includes the foregoing methodwherein the immune disorder is transplant rejection or graft versus hostdisease.

In another embodiment the invention includes a method of using anantibody or antigen binding fragment that specifically binds AI581519_P3(SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13),AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15), AI581519_P10(SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21),AA424839_P7 (SEQ ID NO:23), AA424839_(—)1_P11 (SEQ ID NO:24),H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73), R31375_P33 (SEQ IDNO:74), or a fragment or variant thereof for in vivo imaging of tumorsor inflammatory sites characterized by the differential expression ofAI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15),AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5(SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23), AA424839_(—)1_P11 (SEQ IDNO:24), H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73), R31375_P33 (SEQ IDNO:74), or a fragment or variant thereof.

In another embodiment the invention includes the foregoing method whichis used in assessing cancer prognosis or a treatment protocol.

In another embodiment the invention includes a method for screening fora disease in a subject, comprising detecting in the subject or in asample obtained from said subject a polypeptide having a sequence atleast 85% homologous to the amino acid sequence as set forth in any oneof AI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5(SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15),AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5(SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23), AA424839_(—)1_P11 (SEQ IDNO:24), H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73), R31375_P33 (SEQ IDNO:74), or with a polypeptide having a sequence comprising theextracellular domain of any one of AI581519_P3 (SEQ ID NO:11),AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQID NO:14), AI581519_P9 (SEQ ID NO:15), AI581519_P10 (SEQ ID NO:16),AA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQID NO:23), AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ IDNO:35), H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73), R31375_P33 (SEQ ID NO:74).

In another embodiment the invention includes the foregoing methodwherein screening for a disease comprises detecting the presence orseverity of the disease, disorder or condition, or prognosis of thesubject, or treatment selection for said subject, or treatmentmonitoring of said subject.

In another embodiment the invention includes the foregoing method,wherein the disease is a cancer, selected from the group consisting oflung cancer, ovarian cancer, colon cancer, and wherein the lung cancer,the ovarian cancer and the colon cancer is non-metastatic, invasive ormetastatic.

In another embodiment the invention includes the foregoing methodwherein the disease is autoimmune disease and is selected from multiplesclerosis; psoriasis; rheumatoid arthritis; systemic lupuserythematosus; ulcerative colitis; Crohn's disease; immune disordersassociated with graft transplantation rejection; benign lymphocyticangiitis, lupus erythematosus, Hashimoto's thyroiditis, primarymyxedema, Graves' disease, pernicious anemia, autoimmune atrophicgastritis, Addison's disease, insulin dependent diabetes mellitis, Goodpasture's syndrome, myasthenia gravis, pemphigus, sympatheticophthalmia, autoimmune uveitis, autoimmune hemolytic anemia, idiopathicthrombocytopenia, primary biliary cirrhosis, chronic action hepatitis,ulceratis colitis, Sjogren's syndrome, rheumatic disease, polymyositis,scleroderma, mixed connective tissue disease, inflammatory rheumatism,degenerative rheumatism, extra-articular rheumatism, collagen diseases,chronic polyarthritis, psoriasis arthropathica, ankylosing spondylitis,juvenile rheumatoid arthritis, periarthritis humeroscapularis,panarteriitis nodosa, progressive systemic scleroderma, arthritisuratica, dermatomyositis, muscular rheumatism, myositis, myogelosis andchondrocalcinosis.

In another embodiment the invention includes the foregoing method,wherein the detection is conducted by immunoassay.

In another embodiment the invention includes the foregoing method,wherein the immunoassay utilizes an antibody which specificallyinteracts with the polypeptide having a sequence at least 85% homologousto the amino acid sequence as set forth in any one of AI581519_P3 (SEQID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13),AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15), AI581519_P10(SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21),AA424839_P7 (SEQ ID NO:23), AA424839_(—)1_P11 (SEQ ID NO:24),H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73), R31375_P33 (SEQ IDNO:74), or with a polypeptide having a sequence comprising theextracellular domain of any one of AI581519_P3 (SEQ ID NO:11),AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQID NO:14), AI581519_P9 (SEQ ID NO:15), AI581519_P10 (SEQ ID NO:16),AA424839_P3 (SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQID NO:23), AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ IDNO:35), H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73), and R31375_P33 (SEQ ID NO:74).

In another embodiment the invention includes an antibody specific toAI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5 (SEQID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15),AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5(SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23), AA424839_(—)1_P11 (SEQ IDNO:24), H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73), and R31375_P33(SEQ ID NO:74), or a fragment or variant thereof that elicits apoptosisor lysis of cancer cells that express said protein.

In another embodiment the invention includes any of the foregoingantibodies or fragments, wherein said apoptosis or lysis activityinvolves CDC or ADCC activity of the antibody.

In another embodiment the invention includes any of the foregoingantibodies or fragments, wherein the cancer cells are selected from agroup consisting of hematological malignancies such as acute lymphocyticleukemia, chronic lymphocytic leukemia, acute myelogenous leukemia,chronic myelogenous leukemia, multiple myeloma, Hodgkin's lymphoma,Non-Hodgkin's lymphoma, and soft or solid tumors such as cancer ofbreast, prostate, lung, ovary, colon, spleen, kidney, bladder, head andneck, uterus, testicles, stomach, cervix, liver, bone, skin, pancreas,and brain.

In another embodiment the invention includes any of the foregoingantibodies or fragments, wherein the cancer cells are lung, ovarian orcolon cancer cells.

In another embodiment the invention relates to any of the foregoingisolated soluble VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3ectodomain polypeptides, wherein said polypeptide or a fragment orvariant thereof is used as an anti-cancer vaccine for cancerimmunotherapy.

In another embodiment the invention relates to any an isolatedpolypeptide comprising an amino acid sequence having at least 80%, 85%,90%, 95, 96, 97, 98 or 99%, 100% homologous to the sequence as that setforth in any one of SEQ. ID NOs: 284-295, or a fragment thereof.

In another embodiment the invention relates to any an isolatedpolynucleotide, comprising an amplicon having a nucleic acid sequenceselected from the group consisting of 187, 190, 193, 196, 199, 202, 205,208, 211, 214, 217, 220, 223, 226, 229, 232, 235, 238, 241, 244, 247,250, 253, or polynucleotides homologous thereto.

In another embodiment the invention relates to any a primer pair,comprising a pair of isolated oligonucleotides capable of amplifying theabove mentioned amplicon.

The primer pair, comprising a pair of isolated oligonucleotides having asequence selected from the group consisting of SEQ. ID NOs: 185-186,188-189, 191-192, 194-195, 197-198, 200-201, 203-204, 206-207, 209-210,212-213, 215-216, 218-219, 221-222, 224-225, 227-228, 230-231, 233-234,236-237, 239-240, 242-243, 245-246, 248-249, and 251-252.

A method for screening for a disease, disorder or condition in asubject, comprising detecting in the subject or in a sample obtainedfrom said subject a polynucleotide having a sequence at least 85%homologous to the nucleic acid sequence as set forth in any one of SEQID NOs:1-10, 17-20, 25-34, 41-42, 45-46, 51-69, 187, 190, 193, 196, 199,202, 205, 208, 211, 214, 217, 220, 223, 226, 229, 232, 235, 238, 241,244, 247, 250, and 253.

The method as above, wherein screening for a disease comprises detectingthe presence or severity of the disease, disorder or condition, orprognosis of the subject, or treatment selection for said subject, ortreatment monitoring.

The method as above, wherein the disease is a cancer, selected from thegroup consisting of lung cancer, colon cancer and ovarian cancer, andwherein the lung cancer, colon cancer and ovarian cancer isnon-metastatic, invasive or metastatic.

The method as above, wherein the disease is autoimmune disease.

The method as above, wherein the detection is performed using anoligonucleotide pair capable of hybridizing to at least a portion of anucleic acid sequence at least 85% homologous to the nucleic acidsequence set forth in SEQ. ID NO: 1-10, 17-20, 25-34, 41-42, 45-46,51-69, 187, 190, 193, 196, 199, 202, 205, 208, 211, 214, 217, 220, 223,226, 229, 232, 235, 238, 241, 244, 247, 250, and 253.

The method as above wherein the detection is performed using anoligonucleotide pair as set forth in any one of SEQ. ID NOs: 185-186,188-189, 191-192, 194-195, 197-198, 200-201, 203-204, 206-207, 209-210,212-213, 215-216, 218-219, 221-222, 224-225, 227-228, 230-231, 233-234,236-237, 239-240, 242-243, 245-246, 248-249, and 251-252.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 shows a schematic summary of quantitative real-time PCR analysis.

FIG. 2 shows a scatter plot, demonstrating the expression of AI581519transcripts, that encode the VSIG1 proteins, on a virtual panel of alltissues and conditions using MED discovery engine, demonstratingoverexpression of AI581519 transcripts in lung cancer compared to normallung samples.

FIGS. 3A-3E show alignment comparison of the AI581519_P4 (FIG. 3A),AI581519_P5 (FIG. 3B), AI581519_P7 (FIG. 3C), AI581519_P9 (FIG. 3D), andAI581519_P10 (FIG. 3E) proteins to the known VSIG1 proteins NP_(—)872413(SEQ ID NO: 11) and Q86XK7_HUMAN.

FIG. 4 presents a histogram showing expression of V-set andimmunoglobulin domain containing 1(VSIG1) AI581519 transcripts which aredetectable by amplicon as depicted in sequence name AI581519_seg7 (SEQID NO: 190) in normal and cancerous Ovary tissues.

FIG. 5 presents a histogram showing expression of V-set andimmunoglobulin domain containing 1(VSIG1) AI581519 transcripts which aredetectable by amplicon as depicted in sequence name AI581519_seg7 (SEQID NO: 190) in normal and cancerous lung tissues.

FIGS. 6A-6B present histograms showing expression of V-set andimmunoglobulin domain containing 1(VSIG1) AI581519 transcripts which aredetectable by amplicon as depicted in sequence name AI581519_seg7 (SEQID NO: 190) in various normal tissues. FIG. 6A shows expression of eachsample relative to median of the ovary samples; FIG. 6B shows expressionof each sample relative to median of the lung samples.

FIG. 7 presents a histogram showing expression of V-set andimmunoglobulin domain containing 1(VSIG1) AI581519 transcripts which aredetectable by amplicon as depicted in sequence name AI581519_seg7-9 (SEQID NO: 187) in normal and cancerous Ovary tissues.

FIG. 8 presents a histogram showing expression of V-set andimmunoglobulin domain containing 1(VSIG1) AI581519 transcripts which aredetectable by amplicon as depicted in sequence name AI581519_seg7-9 (SEQID NO: 187) in normal and cancerous lung tissues.

FIG. 9 presents a histogram showing expression of V-set andimmunoglobulin domain containing 1(VSIG1) AI581519 transcripts which aredetectable by amplicon as depicted in sequence name AI581519seg7-9 (SEQID NO: 196) in blood-specific panel.

FIG. 10 presents a histogram showing expression of V-set andimmunoglobulin domain containing 1(VSIG1) AI581519 transcripts which aredetectable by amplicon as depicted in sequence nameAI581519_junc7-11F2R2 (SEQ ID NO: 193) in normal and cancerous lungtissues.

FIG. 11 presents a histogram showing expression of V-set andimmunoglobulin domain containing 1(VSIG1) AI581519 transcripts which aredetectable by amplicon as depicted in sequence nameAI581519_junc7-11F2R2 (SEQ ID NO: 193) in normal and cancerous ovariantissues.

FIGS. 12A-12B present histograms showing expression of V-set andimmunoglobulin domain containing 1(VSIG1) AI581519 transcripts which aredetectable by amplicon as depicted in sequence nameAI581519_junc7-11F2R2 (SEQ ID NO: 193) in various normal tissues. FIG.12A shows expression of each sample relative to median of the lungsamples; FIG. 12B shows expression of each sample relative to median ofthe ovary samples.

FIG. 13 presents a histogram showing expression of V-set andimmunoglobulin domain containing 1(VSIG1) AI581519 transcripts which aredetectable by amplicon as depicted in sequence nameAI581519_junc7-11F2R2 (SEQ ID NO: 193) in blood-specific panel.

FIG. 14 shows a scatter plot, demonstrating the expression of AA424839transcripts, that encode the ILDR1 proteins, on a virtual panel of alltissues and conditions using MED discovery engine, demonstratingoverexpression of AA424839 transcripts in ovarian cancer compared tonormal ovary samples.

FIG. 15 shows a scatter plot, demonstrating the expression of AA424839transcripts, that encode the ILDR1 proteins, on a virtual panel of alltissues and conditions using MED discovery engine, demonstratingoverexpression of AA424839 transcripts in colon cancer compared tonormal colon samples.

FIGS. 16A-16C show alignment comparison of the AA424839_P3 (FIG. 16A),AA424839_P7 (FIG. 16B), and AA424839_(—)1_P11 (FIG. 16C), proteins tothe known ILDR1 proteins Q86SU0_HUMAN and NP_(—)787120 (SEQ ID NO: 21).

FIG. 17 presents a histogram showing expression of immunoglobulin-likedomain containing receptor 1 (ILDR1) AA424839 transcripts which aredetectable by amplicon as depicted in sequence name AA424839_seg18wt(SEQ ID NO: 199) in normal and cancerous ovary tissues.

FIG. 18 presents a histogram showing expression of immunoglobulin-likedomain containing receptor 1 (ILDR1) AA424839 transcripts which aredetectable by amplicon as depicted in sequence name AA424839_seg18wt(SEQ ID NO: 199) in various normal tissues.

FIG. 19 presents a histogram showing expression of immunoglobulin-likedomain containing receptor 1 (ILDR1) AA424839 transcripts which aredetectable by amplicon as depicted in sequence name AA424839_seg14-16(SEQ ID NO: 202) in normal and cancerous ovary tissues.

FIG. 20 presents a histogram showing expression of immunoglobulin-likedomain containing receptor 1 (ILDR1) AA424839 transcripts which aredetectable by amplicon as depicted in sequence name AA424839_seg14-16(SEQ ID NO: 202) in various normal tissues.

FIG. 21 presents a histogram showing expression of immunoglobulin-likedomain containing receptor 1 (ILDR1) AA424839 transcripts which aredetectable by amplicon as depicted in sequence name AA424839seg11-14F3R3(SEQ ID NO: 205) in blood-specific panel.

FIG. 22 presents a histogram showing expression of AI216611 transcriptswhich are detectable by amplicon as depicted in sequence nameAI216611_junc4-6F2R2 (SEQ ID NO: 208) in normal and cancerous colontissues.

FIG. 23 presents a histogram showing expression of AI216611 transcriptswhich are detectable by amplicon as depicted in sequence nameAI216611_junc4-6F2R2 (SEQ ID NO: 208) in various normal tissues.

FIG. 24 presents a histogram showing expression of AI216611 transcriptswhich are detectable by amplicon as depicted in sequence nameAI216611_seg2WT (SEQ ID NO: 211) in normal and cancerous colon tissues.

FIG. 25 presents a histogram showing expression of AI216611 transcriptswhich are detectable by amplicon as depicted in sequence nameAI216611_seg2WT (SEQ ID NO: 211) in various normal tissues.

FIG. 26 presents a histogram showing expression of AI216611 transcriptswhich are detectable by amplicon as depicted in sequence nameAI216611junc4-6 (SEQ ID NO: 214) in blood-specific pane.

FIG. 27 presents a histogram showing expression of AI216611 transcriptswhich are detectable by amplicon as depicted in sequence nameAI216611_junc2-4seg5F2R2 (SEQ ID NO: 220) in blood-specific pane.

FIG. 28 presents a histogram showing expression of AI216611 transcriptswhich are detectable by amplicon as depicted in sequence nameAI216611_junc2-4seg5F2R2 (SEQ ID NO: 220) in normal and cancerous colontissues.

FIG. 29 shows a scatter plot, demonstrating the expression of H68654transcripts, that encode the LOC253012 proteins, on a virtual panel ofall tissues and conditions using MED discovery engine, demonstratingoverexpression of H68654 transcripts in lung cancer compared to normallung samples.

FIGS. 30A-30H show alignment comparison of the H68654_(—)1_P7 (FIG. 30Aand FIG. 30B), H68654_(—)1_P12 (FIGS. 3C and 30D), H68654_(—)1_P13 (FIG.3E and FIG. 30F), and H68654_(—)1_P14 (FIGS. 30G and 30H) proteins tothe known LOC253012 proteins NP_(—)937794 and Q6UXI0_HUMAN (SEQ ID NO:36).

FIG. 31 presents a histogram showing expression of hypothetical proteinLOC253012 H68654 transcripts which are detectable by amplicon asdepicted in sequence name H68654_seg3WTF2R2 (SEQ ID NO: 226) in normaland cancerous Lung tissues.

FIG. 32 presents a histogram showing expression of hypothetical proteinLOC253012 H68654 transcripts which are detectable by amplicon asdepicted in sequence name H68654_seg3WTF2R2 (SEQ ID NO: 226) in variousnormal and tissues.

FIG. 33 presents a histogram showing expression of hypothetical proteinLOC253012 H68654 transcripts which are detectable by amplicon asdepicted in sequence name H68654_seg7-12WT (SEQ ID NO: 223) in normaland cancerous Lung tissues.

FIG. 34 presents a histogram showing expression of hypothetical proteinLOC253012 H68654 transcripts which are detectable by amplicon asdepicted in sequence name H68654_seg7-12WT (SEQ ID NO: 223) in variousnormal and tissues.

FIG. 35A presents a histogram showing expression of hypothetical proteinLOC253012 H68654 transcripts which are detectable by amplicon asdepicted in sequence name H68654seg3F2R2 (SEQ ID NO: 226) inblood-specific panel.

FIG. 35B presents a histogram showing expression of hypothetical proteinLOC253012 H68654 transcripts which are detectable by amplicon asdepicted in sequence name H68654seg7-12F1R1 (SEQ ID NO: 223) inblood-specific panel.

FIG. 36 presents a histogram showing expression of hypothetical proteinLOC253012 H68654 transcripts which are detectable by amplicon asdepicted in sequence name H68654_seg0-3 (SEQ ID NO: 229) in normal andcancerous Lung tissues.

FIG. 37 presents a histogram showing expression of hypothetical proteinLOC253012 H68654 transcripts which are detectable by amplicon asdepicted in sequence name H68654_seg2-3 (SEQ ID NO: 232) in normal andcancerous Lung tissues.

FIGS. 38A-38B show alignment comparison of the H19011_(—)1_P8 (FIG. 38A)and H19011_(—)1_P9 (FIG. 38B) proteins to the known C1ORF32 proteinsQ71H61_HUMAN and NP_(—)955383 (SEQ ID NO: 47).

FIG. 39 presents a histogram showing expression of C1ORF32, chromosome 1open reading frame 32, H19011 transcripts which are detectable byamplicon as depicted in sequence name H19011_seg13F2R2 (SEQ ID NO: 235)in normal and cancerous Colon tissues.

FIG. 40 presents a histogram showing expression of C1ORF32, chromosome 1open reading frame 32, H19011 transcripts which are detectable byamplicon as depicted in sequence name H19011_seg13F2R2 (SEQ ID NO: 235)in normal and cancerous lung tissues.

FIGS. 41A-41B present a histogram showing expression of C1ORF32,chromosome 1 open reading frame 32, H19011 transcripts which aredetectable by amplicon as depicted in sequence name H19011_seg13F2R2(SEQ ID NO: 235) in various normal tissues. FIG. 41A shows expression ofeach sample relative to median of the colon samples; FIG. 41B showsexpression of each sample relative to median of the lung samples.

FIG. 42 presents a histogram showing expression of C1ORF32, chromosome 1open reading frame 32, H19011 transcripts which are detectable byamplicon as depicted in sequence name H19011_seg8-13F1R1 (SEQ ID NO:238) in normal and cancerous lung tissues.

FIG. 43 presents a histogram showing expression of C1ORF32, chromosome 1open reading frame 32, H19011 transcripts which are detectable byamplicon as depicted in sequence name H19011_junc8-10seg13 (SEQ ID NO:241) in normal and cancerous lung tissues.

FIG. 44 presents a histogram showing expression of C1ORF32, chromosome 1open reading frame 32, H19011 transcripts which are detectable byamplicon as depicted in sequence name H19011_junc8-10seg13 (SEQ ID NO:241) in normal and cancerous colon tissues.

FIGS. 45A-45B presents a histogram showing expression of C1ORF32,chromosome 1 open reading frame 32, H19011 transcripts which aredetectable by amplicon as depicted in sequence name H19011_junc8-10seg13(SEQ ID NO: 241) in various normal tissues. FIG. 45A shows expression ofeach sample relative to median of the colon samples; FIG. 45B showsexpression of each sample relative to median of the lung samples.

FIG. 46 presents a histogram showing expression of C1ORF32, chromosome 1open reading frame 32, H19011 transcripts which are detectable byamplicon as depicted in sequence name H19011_junc8-10seg13 (SEQ ID NO:241) in blood-specific panel.

FIG. 47 presents a histogram showing expression of C1ORF32, chromosome 1open reading frame 32, H19011 transcripts which are detectable byamplicon as depicted in sequence name H19011_junc6-10F1R1 (SEQ ID NO:244) in normal and cancerous lung tissues.

FIG. 48 presents a histogram showing expression of C1ORF32, chromosome 1open reading frame 32, H19011 transcripts which are detectable byamplicon as depicted in sequence name H19011_junc6-10F1R1 (SEQ ID NO:244) in normal and cancerous colon tissues.

FIGS. 49A-49E show alignment comparison of the R31375_P14 (FIG. 49A),R31375_P31 (FIGS. 49B and 49C) and R31375_P33 (FIGS. 49D and 49E),proteins to the known FXYD3 proteins NP_(—)068710 (SEQ ID NO: 71),FXYD3_HUMAN (SEQ ID NO: 70), NP_(—)005962 and Q6IB59_HUMAN (SEQ ID NO:70).

FIG. 50 presents a histogram showing expression of FXYD3 domaincontaining ion transport regulator 3 R31375 transcripts which aredetectable by amplicon as depicted in sequence name R31375_junc30-33(SEQ ID NO: 247) in normal and cancerous ovary tissues.

FIG. 51 presents a histogram showing expression of FXYD3 domaincontaining ion transport regulator 3 R31375 transcripts which aredetectable by amplicon as depicted in sequence name R31375_junc30-33(SEQ ID NO: 247) in various normal tissues.

FIG. 52 presents a histogram showing expression of FXYD3 domaincontaining ion transport regulator 3 R31375 transcripts which aredetectable by amplicon as depicted in sequence nameR31375_seg33junc34-37 (SEQ ID NO: 250) in normal and cancerous ovarytissues.

FIG. 53 presents a histogram showing expression of FXYD3 domaincontaining ion transport regulator 3 R31375 transcripts which aredetectable by amplicon as depicted in sequence nameR31375_seg33junc34-37 (SEQ ID NO: 250) in various normal tissues.

FIG. 54 presents a histogram showing expression of FXYD3 domaincontaining ion transport regulator 3 R31375 transcripts which aredetectable by amplicon as depicted in sequence nameR31375_junc20-22seg30F6R6 (SEQ ID NO: 253) in normal and cancerous ovarytissues.

FIG. 55 presents a histogram showing expression of FXYD3 domaincontaining ion transport regulator 3 R31375 transcripts which aredetectable by amplicon as depicted in sequence nameR31375_junc20-22seg30F6R6 (SEQ ID NO: 253) in various normal tissues.

FIG. 56A-56J presents the nucleotide sequences of the recombinant fulllength_EGFP ORFs: gene specific sequence correspond to the candidate'sfull length sequence is marked in bold, EGFP sequence is unbold Italicand known SNPs/silence mutation are underlined. FIG. 56A presents thefull length_EGFP ORF nucleic acid sequence of FXYD3_T0_P0_EGFP DNA (996bp) (SEQ ID NO:77); FIG. 56B presents the full length_EGFP ORF nucleicacid sequence of FXYD3_T25_P14_EGFP DNA (1083 bp) (SEQ ID NO:78); FIG.56C presents the full length_EGFP ORF nucleic acid sequence ofAI216611_T0_P0_EGFP DNA (1371 bp) (SEQ ID NO:79); FIG. 56D presents thefull length_EGFP ORF nucleic acid sequence of AI216611_T1_P1_EGFP DNA(1332 bp) (SEQ ID NO:80); FIG. 56E presents the full length_EGFP ORFnucleic acid sequence of C1ORF32_T8_P8_EGFP DNA (1533 bp) (SEQ IDNO:81); FIG. 56F presents the full length_EGFP ORF nucleic acid sequenceof LOC253012_T4_P5_EGFP DNA (2085 bp) (SEQ ID NO:82); FIG. 56G presentsthe full length_EGFP ORF nucleic acid sequence of ILDR1_T0_P3_EGFP DNA(2373 bp) (SEQ ID NO:83); FIG. 56H presents the full length_EGFP ORFnucleic acid sequence of ILDR1_T2_P5_EGFP DNA (2241 bp) (SEQ ID NO:84);FIG. 56I presents the full length_EGFP ORF nucleic acid sequence ofVSIG1_T6_P5_EGFP DNA (2082 bp) (SEQ ID NO:85); FIG. 56J presents thefull length_EGFP ORF nucleic acid sequence of VSIG1_T5_P4_EGFP DNA (2004bp) (SEQ ID NO:86).

FIG. 57A-57J presents the sequences of the full length_EGFP fusionproteins of invention. Candidate's specific sequence corresponding tothe full length sequence of the protein is marked in bold, EGFP sequenceis unbold Italic and amino acids modified due to known SNPs areunderlined. FIG. 57A presents the full length_EGFP ORF amino acidsequence of FXYD3_P0_EGFP protein (331aa) (SEQ ID NO:87); FIG. 57Bpresents the full length_EGFP ORF amino acid sequence of FXYD3_P14_EGFPprotein (360aa) (SEQ ID NO:88); FIG. 57C presents the full length_EGFPORF amino acid sequence of AI216611_P0_EGFP protein (456aa) (SEQ IDNO:89); FIG. 57D presents the full length_EGFP ORF amino acid sequenceof AI216611_P1_EGFP protein (443aa) (SEQ ID NO:90); FIG. 57E presentsthe full length_EGFP ORF amino acid sequence of C1ORF32_P8_EGFP protein(510aa) (SEQ ID NO:91); FIG. 57F presents the full length_EGFP ORF aminoacid sequence of LOC253012_P5_EGFP protein (694aa) (SEQ ID NO:92); FIG.57G presents the full length_EGFP ORF amino acid sequence ofILDR1_P3_EGFP protein (790aa) (SEQ ID NO:93); FIG. 57H presents the fulllength_EGFP ORF amino acid sequence of ILDR1_P5_EGFP protein (746aa)(SEQ ID NO:94); FIG. 57I presents the full length_EGFP ORF amino acidsequence of VSIG1_P5_EGFP protein (693aa) (SEQ ID NO:95); FIG. 57Jpresents the full length_EGFP ORF amino acid sequence of VSIG1_P4_EGFPprotein (667aa) (SEQ ID NO:96).

FIGS. 58A-58F demonstrate the localization of the proteins of inventionto cell membrane: FIG. 58A shows cellular localization ofAI216611-EGFP_T0_P0 and AI216611-EGFP_T1_P1 proteins FIG. 58B showscellular localization of FXYD3-EGFP_T0_P0 and FXYD3-EGFP_T25_P14proteins. FIG. 58C shows cellular localization of C1ORF32-EGFP_T8_P8protein. FIG. 58D shows cellular localization of LOC253012-EGFP_T4_P5protein. FIG. 58E shows cellular localization of VSIG1-EGFP_T6_P5 andVSIG1-EGFP_T5_P4 proteins. FIG. 58F shows cellular localization ofILDR1_EGFP_T0_P3 and ILDR1_EGFP_T2_P5 proteins. All the images wereobtained using the 40× objective of the confocal microscope.

FIGS. 59A-59F present the nucleotide sequences of the extracellulardomains of the candidate proteins of the invention, fused to mouse Fc:ECD_mFc ORFs. Candidate protein's specific sequence corresponding to theECD sequence is marked in bold, TEV cleavage site sequence isunderlined, mFc sequence is unbold Italic and IL6sp sequence is boldItalic. FIG. 59A shows the FXYD3_T25_P14_ECD_mFc DNA sequence (924 bp)(SEQ ID NO:97); FIG. 59B shows the AI216611_T0_P0_ECD_mFc DNA sequence(1170 bp) (SEQ ID NO:98), FIG. 59C shows the C1ORF32_T8_P8_ECD_mFc DNAsequence (1287 bp) (SEQ ID NO:99); FIG. 59D shows theLOC253012_T4_P5_ECD_mFc DNA sequence (1740 bp) (SEQ ID NO:100), FIG. 59Eshows the ILDR1_T0_P3_ECD_mFc DNA sequence (1167 bp) (SEQ ID NO:101),and FIG. 59F shows the VSIG1_T6_P5_ECD_mFc DNA sequence (1641 bp) (SEQID NO:102).

FIGS. 60A-60F present the amino acid sequence of the ECD_mFc fusionproteins. Candidate protein's specific sequence corresponding to the ECDsequence is marked in bold, TEV cleavage site sequence is underlined,mFc sequence is unbold Italic and IL6sp sequence is bold Italic. FIG.60A shows the FXYD3_T25_P14_ECD_mFc amino acid sequence (307aa) (SEQ IDNO:103); FIG. 60B shows the AI216611_T0_P0_ECD_mFc amino acid sequence(389aa) (SEQ ID NO:104), FIG. 60C shows the C1ORF32_T8_P8_ECD_mFc aminoacid sequence (428aa) (SEQ ID NO:105); FIG. 60D shows theLOC253012_T4_P5_ECD_mFc amino acid sequence (579aa) (SEQ ID NO:106),FIG. 60E shows the ILDR1_T0_P3_ECD_mFc amino acid sequence (388aa) (SEQID NO:107), and FIG. 60F shows the VSIG1_T6_P5_ECD_mFc amino acidsequence (546aa) (SEQ ID NO:108).

FIG. 61 shows the results of a western blot analysis of the expressedFXYD3_ECD_mFc (SEQ ID NO:103), AI216611 ECD_mFc (SEQ ID NO:104),C1ORF32_ECD_mFc (SEQ ID NO:105), LOC253012_ECD_mFc (SEQ ID NO:106),ILDR1_ECD_mFc (SEQ ID NO:107), VSIG1_ECD_mFc (SEQ ID NO:108). The lanesare as follows: lane 1 Molecular weight markers (Amersham, full rangeranbow, catalog number RPN800); lane 2—LOC253012_ECD_mFc; lane3—FXYD3_ECD_mFc; lane 4—AI216611 ECD_mFc; lane 5—C1ORF32_ECD_mFc; lane6—ILDR1_ECD_mFc; lane 7—VSIG1_ECD_mFc.

FIGS. 62A-62E present the binding of the Fc-fused B7-like proteins ECDsto resting T cells or T cells activated with Con A for different periodsof time. FIG. 62A shows the binding results for Fc-fused VSIG1 ECD; FIG.62B shows the binding results for Fc-fused LOC253012; FIG. 62C shows thebinding results for Fc-fused C1ORF32 ECD; and FIG. 62D shows the bindingresults for Fc-fused AI216611 ECD. FIG. 62E shows the binding resultsfor Fc-fused FXYD3 ECD.

FIG. 63 presents the dose response of the binding of Fc-fused B7-likeproteins ECDs to activated T cells. Purified T cells were cultured for48 hours. Con A was added for the last 24 hours. Cells were thenharvested and stained with increasing concentrations (3, 6, 12, 25 and50 μg/ml) of Fc-fused VSIG1, LOC253012, C1ORF32, AI216611, ILDR1 orFXYD3 ECDs. As negative controls, mouse IgG2a was used at the sameconcentrations.

FIGS. 64A-64B present the effect of the ECD-Fc fused proteins on T cellsproliferation or IL-2 secretion, upon activation with anti-CD3 Ab. FIG.64A shows the levels of BrdU incorporation. FIG. 64B shows the levels ofIL-2 secretion.

FIG. 65 illustrates the binding of the Fc-fused ECDs of the VSIG1,ILDR1, LOC253012, AI216611, FXYD3 or C1ORF32 to lymphocytes.

FIG. 66 illustrates the binding of the Fc-fused ECDs of the ILDR1,C1ORF32 and AI216611 to CD4+ T cells.

FIG. 67 shows the effect of B7-like proteins on T cell activation. “CD3”means CD3 only without the presence of a costimulatory molecule;“CD3+B7.2” means CD3+a known B7 stimulatory control, B7.2; “CD3+B7H4”means CD3 and B7H4 a known B7 inhibitory control; “CD3+B7H3” means CD3and B7H3 a known B7 stimulatory protein; “CD3+702” meansCD3+LOC253012-ECD-Fc fused (SEQ ID NO:106); “CD3+721” meansCD3+AI216611-ECD-Fc fused (SEQ ID NO:104); “CD3+754” meansCD3+C1ORF32-ECD-Fc fused (SEQ ID NO:105); “CD3+768” meansCD3+VSIG1-ECD-Fc fused (SEQ ID NO:108) “CD3+770” means CD3+ILDR1_ECD-Fcfused (SEQ ID NO:107); “CD3+789” means CD3+FXYD3-ECD-Fc fused (SEQ IDNO:103). FIGS. 67A, B and C present 3 different experiments of 3different donors

FIG. 68A presents FACS results of binding of ILDR1_ECD-Fc (SEQ IDNO:107), C1ORF32-ECD-Fc (SEQ ID NO:105), AI216611-ECD-Fc (SEQ IDNO:104), LOC253012-ECD-Fc (SEQ ID NO:106), FXYD3-ECD-Fc (SEQ ID NO:103),and VSIG1-ECD-Fc (SEQ ID NO:108) to resting B cells

FIG. 68B presents FACS results of binding of binding of ILDR1_ECD-Fc(SEQ ID NO:107), C1ORF32-ECD-Fc (SEQ ID NO:105), AI216611-ECD-Fc (SEQ IDNO:104), LOC253012-ECD-Fc (SEQ ID NO:106), FXYD3-ECD-Fc (SEQ ID NO:103),and VSIG1-ECD-Fc (SEQ ID NO:108) to activated B cells.

FIG. 68C presents FACS results of binding of ILDR1_ECD-Fc (SEQ IDNO:107), C1ORF32-ECD-Fc (SEQ ID NO:105), AI216611-ECD-Fc (SEQ IDNO:104), LOC253012-ECD-Fc (SEQ ID NO:106), FXYD3-ECD-Fc (SEQ ID NO:103),and VSIG1-ECD-Fc (SEQ ID NO:108) to B lymphoma cell lines.

FIG. 69 shows BIACORE results demonstrating interaction between AI216611and B7H4.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to any one of the antigens referred to asVSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3, and its correspondingnucleic acid sequence, and portions and variants thereof and conjugatescontaining and the use thereof as a therapeutic or diagnostic target. Inparticular the invention uses this antigen and discrete portions thereofas a drug target for therapeutic small molecules, peptides, antibodies,antisense RNAs, siRNAs, ribozymes, and the like. More particularly theinvention relates to diagnostic and therapeutic polyclonal andmonoclonal antibodies and fragments thereof that bind VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, FXYD3 and portions and variants thereof,especially those that target the ectodomain or portions or variantsthereof particularly human or chimeric monoclonal antibodies, that bindspecifically to the antigen AI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQID NO:12), AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14),AI581519_P9 (SEQ ID NO:15), AI581519_P10 (SEQ ID NO:16), AA424839_P3(SEQ ID NO:22), AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23),AA424839_(—)1_P11 (SEQ ID NO:24), H68654_(—)1_P2 (SEQ ID NO:35),H68654_(—)1_P5 (SEQ ID NO:36), H68654_(—)1_P7 (SEQ ID NO:37),H68654_(—)1_P12 (SEQ ID NO:38), H68654_(—)1_P13 (SEQ ID NO:39),H68654_(—)1_P14 (SEQ ID NO:40), AI216611_P0 (SEQ ID NO:43), AI216611_P1(SEQ ID NO:44), H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72), R31375_P31(SEQ ID NO:73), R31375_P33 (SEQ ID NO:74), and variants thereofincluding those that promote or inhibit activities elicited by VSIG1,ILDR1, LOC253012, AI216611, C1ORF32, FXYD3, including those relating tomodulation of immune costimulation, e.g. B7 related costimulation.

In certain embodiments, the antibodies of the invention are derived fromparticular heavy and light chain germline sequences and/or compriseparticular structural features such as CDR regions comprising particularamino acid sequences. The invention provides isolated antibodies,methods of making such antibodies, immunoconjugates and bispecificmolecules comprising such antibodies and pharmaceutical and diagnosticcompositions containing the antibodies, immunoconjugates or bispecificmolecules of the invention.

The invention also relates to in vitro and in vivo methods of using theantibodies and fragments, to detect VSIG1, ILDR1, LOC253012, AI216611,C1ORF32, FXYD3, as well as to treat diseases associated with expressionof VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3, such asmalignancies that differentially express VSIG1. The invention furtherrelates to methods of using the antibodies and fragments, specific forVSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 to treat autoimmunedisorders and transplant and graft versus host disease. Accordingly, theinvention also provides methods of using the anti-VSIG1, anti-ILDR1,anti-LOC253012, anti-AI216611, anti-C1ORF32, anti-FXYD3 antibodies ofthe invention and other drugs that modulate VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, FXYD3 to treat malignancies for example, in thetreatment of lung cancer, ovarian cancer, colon cancer, non-solid andsolid tumors, sarcomas, hematological malignancies including but notlimited to acute lymphocytic leukemia, chronic lymphocytic leukemia,acute myelogenous leukemia, chronic myelogenous leukemia, multiplemyeloma, Hodgkin's lymphoma, Non-Hodgkin's lymphoma, cancer of thebreast, prostate, spleen, kidney, bladder, head and neck, uterus,testicles, stomach, cervix, liver, bone, skin, pancreas, brain andwherein the cancer may be non-metastatic, invasive or metastatic. Theinvention further provides methods of using the anti-VSIG1, anti-ILDR1,anti-LOC253012, anti-AI216611, anti-C1ORF32, anti-FXYD3 antibodies ofthe invention and other drugs that modulate VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, FXYD3 to treat non-malignant disorders such as immunedisorders including but not limited to autoimmune diseases, transplantrejection and graft versus host disease. Preferably these antibodieswill possess ADCC or CDC activity against target cells such as cancercells.

Also, the invention relates to the VSIG1, ILDR1, LOC253012, AI216611,C1ORF32, FXYD3 antigen and portions thereof including solublepolypeptide conjugates containing the ectodomain of VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, FXYD3 and/or the corresponding DNAs orvectors or cells expressing same for use in immunotherapy. Further theinvention provides vectors, cells containing and use thereof for theexpression of the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3antigen, as well as discrete portions and variants thereof. Also, theinvention provides non-antibody based VSIG1, ILDR1, LOC253012, AI216611,C1ORF32, FXYD3 modulatory agents such as peptides, antisense RNAs,siRNAs, carbohydrates, and other small molecules that specifically bindand/or modulate a VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3related activity.

In order that the present invention may be more readily understood,certain terms are first defined. Additional definitions are set forththroughout the detailed description.

The terms VSIG1 refers to the protein encoded by any one of theAI581519_TO (SEQ ID NO:1), AI581519_T1 (SEQ ID NO:2), AI581519_T2 (SEQID NO:3), AI581519_T3 (SEQ ID NO:4), AI581519_T4 (SEQ ID NO:5),AI581519_T5 (SEQ ID NO:6), AI581519_T6 (SEQ ID NO:7), AI581519_T8 (SEQID NO:8), AI581519_T10 (SEQ ID NO:9), AI581519_T11 (SEQ ID NO:10)transcripts reported herein, particularly to proteins as set forth inany one of AI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12),AI581519_P5 (SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQID NO:15), AI581519_P10 (SEQ ID NO:16), and variants thereof that aredifferentially expressed e.g., in cancers such as lung cancer andovarian cancer, wherein the cancer may be non-metastatic, invasive ormetastatic as well as non-malignant disorders such as immune disordersincluding but not limited to autoimmune diseases, transplant rejectionand graft versus host disease.

The term ILDR1 refers to the to the protein encoded by any one of theAA424839_TO (SEQ ID NO:17), AA424839_T2 (SEQ ID NO:18), AA424839_T4 (SEQID NO:19), AA424839_(—)1_T7 (SEQ ID NO:20) transcripts reported herein,particularly to proteins as set forth in any one of AA424839_P3 (SEQ IDNO:22), AA424839_P5 (SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23),AA424839_(—)1_P11 (SEQ ID NO:24), and variants thereof that aredifferentially expressed e.g., in cancer such as colon cancer andovarian cancer wherein the cancer may be non-metastatic, invasive ormetastatic as well as non-malignant disorders such as immune disordersincluding but not limited to autoimmune diseases, transplant rejectionand graft versus host disease.

The term LOC253012 refers to the protein encoded by any one of theH68654_(—)1_TO (SEQ ID NO:25), H68654_(—)1_T4 (SEQ ID NO:26),H68654_(—)1_T5 (SEQ ID NO:27), H68654_(—)1_T8 (SEQ ID NO:28),H68654_(—)1_T15 (SEQ ID NO:29), H68654_(—)1_T16 (SEQ ID NO:30),H68654_(—)1_T17 (SEQ ID NO:31), H68654_(—)1_T18 (SEQ ID NO:32),H68654_(—)1_T19 (SEQ ID NO:33), H68654_(—)1_T20 (SEQ ID NO:34)transcripts reported herein, particularly to proteins as set forth inany one of H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40), andvariants thereof that are differentially expressed e.g., in cancers suchas lung cancer, especially small cell lung carcinoma, wherein the cancermay be non-metastatic, invasive or metastatic as well as non-malignantdisorders such as immune disorders including but not limited toautoimmune diseases, transplant rejection and graft versus host disease.

The term AI216611 refers to the protein encoded by any one of theAI216611_TO (SEQ ID NO:41), AI216611_T1 (SEQ ID NO:42) transcriptsreported herein, particularly to proteins as set forth in any one ofAI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), and variantsthereof that are differentially expressed e.g., in cancers such asnon-solid and solid tumors, sarcomas, hematological malignanciesincluding but not limited to acute lymphocytic leukemia, chroniclymphocytic leukemia, acute myelogenous leukemia, chronic myelogenousleukemia, multiple myeloma, Hodgkin's lymphoma, Non-Hodgkin's lymphoma,cancer of the breast, prostate, lung, ovary, colon, spleen, kidney,bladder, head and neck, uterus, testicles, stomach, cervix, liver, bone,skin, pancreas, and brain and wherein the cancer may be non-metastatic,invasive or metastatic as well as non-malignant disorders such as immunedisorders including but not limited to autoimmune diseases, transplantrejection and graft versus host disease.

The terms C1ORF32 refers to the protein encoded by any one of theH19011_(—)1_T8 (SEQ ID NO:45), H19011_(—)1_T9 (SEQ ID NO:46) transcriptsreported herein, particularly to proteins as set forth in any one ofH19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), andvariants thereof that are differentially expressed e.g., in cancers suchas lung cancer, particularly lung small cell carcinoma, wherein thecancer may be non-metastatic, invasive or metastatic as well asnon-malignant disorders such as immune disorders including but notlimited to autoimmune diseases, transplant rejection and graft versushost disease.

The term FXYD3 refers to the protein encoded by any one of the R31375_TO(SEQ ID NO:51); R31375_T1 (SEQ ID NO:52); R31375_T10 (SEQ ID NO:61);R31375_T11 (SEQ ID NO:62); R31375_T12 (SEQ ID NO:63); R31375_T13 (SEQ IDNO:64); R31375_T2 (SEQ ID NO:53); R31375_T3 (SEQ ID NO:54); R31375_T4(SEQ ID NO:55); R31375_T5 (SEQ ID NO:56); R31375_T6 (SEQ ID NO:57);R31375_T7 (SEQ ID NO:58); R31375_T8 (SEQ ID NO:59); R31375_T9 (SEQ IDNO:60): R31375_T19 (SEQ ID NO:65); R31375_T25 (SEQ ID NO:66); R31375_T26(SEQ ID NO:67); R31375_T29 (SEQ ID NO:68); R31375_T39 (SEQ ID NO:69)transcripts reported herein, particularly to proteins as set forth inany one of R31375_P0 (SEQ ID NO:70), R31375_P14 (SEQ ID NO:72),R31375_P31 (SEQ ID NO:73), R31375_P31 (SEQ ID NO:73), and variantsthereof that are differentially expressed e.g., in cancers such asovarian cancer as well as other non-solid and solid tumors, sarcomas,hematological malignancies including but not limited to acutelymphocytic leukemia, chronic lymphocytic leukemia, acute myelogenousleukemia, chronic myelogenous leukemia, multiple myeloma, Hodgkin'slymphoma, Non-Hodgkin's lymphoma, cancer of the breast, prostate, lung,ovary, colon, spleen, kidney, bladder, head and neck, uterus, testicles,stomach, cervix, liver, bone, skin, pancreas, brain and wherein thecancer may be non-metastatic, invasive or metastatic.

Preferably such VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3variants will possess at least 80% sequence identity therewith, morepreferably at least 90% sequence identity therewith and even morepreferably at least 95% sequence identity therewith.

Any one of the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 proteins basedon its domain structure is predicted to be an immune costimulatoryprotein, e.g., a B7 protein family member that is involved in B7 immuneco-stimulation including for example T cell responses elicited againstcancer cells and that elicit effects on immunity such as triggering ofautoimmune effects.

The term the “soluble ectodomain (ECD)” or “ectodomain” of VSIG1 refersto the polypeptide sequences below or the corresponding nucleic acidsequences (which does not comprise the signal peptide and the TM ofVSIG1 protein):

>AI581519_P3 (SEQ IDS NO: 11) residues 23 to 234 (SEQ ID NO: 138)QVTIPDGFVNVTVGSNVTLICIYTTTVASREQLSIQWSFFHKKEMEPISIYFSQGGQAVAIGQFKDRITGSNDPGNASITISHMQPADSGIYICDVNNPPDFLGQNQGILNVSVLVKPSKPLCSVQGRPETGHTISLSCLSALGTPSPVYYWHKLEGRDIVPVKENFNPTTGILVIGNLTNFEQGYYQCTAINRLGNSSCEIDLTSSHPEVG >AI581519_P4 (SEQ IDS NO: 12) residues 23 to 270 (SEQ ID NO: 139)QVTIPDGFVNVTVGSNVTLICIYTTTVASREQLSIQWSFFHKKEMEPISHSSCLSTEGMEEKAVGQCLKMTHVRDARGRCSWTSEIYFSQGGQAVAIGQFKDRITGSNDPGNASITISHMQPADSGIYICDVNNPPDFLGQNQGILNVSVLVKPSKPLCSVQGRPETGHTISLSCLSALGTPSPVYYWHKLEGRDIVPVKENFNPTTGILVIGNLTNFEQGYYQCTAINRLGNSSCEIDLTSSHPEVG >AI581519_P5 (SEQ IDS NO: 13) residues 23 to 296 (SEQ ID NO: 140)QVTIPDGFVNVTVGSNVTLICIYTTTVASREQLSIQWSFFHKKEMEPISHSSCLSTEGMEEKAVGQCLKMTHVRDARGRCSWTSESPWEEGKWPDVEAVKGTLDGQQAELQIYFSQGGQAVAIGQFKDRITGSNDPGNASITISHMQPADSGIYICDVNNPPDFLGQNQGILNVSVLVKPSKPLCSVQGRPETGHTISLSCLSALGTPSPVYYWHKLEGRDIVPVKENFNPTTGILVIGNLTNFEQGYYQCTAINRLGNSSCEIDLTSSHPEVG >AI581519_P7 (SEQ IDS NO: 14) residues 23 To 193 (SEQ ID NO: 141)QVTIPDGFVNVTVGSNVTLICIYTTTVASREQLSIQWSFFHKKEMEPISIYFSQGGQAVAIGQFKDRITGSNDPVKPSKPLCSVQGRPETGHTISLSCLSALGTPSPVYYWHKLEGRDIVPVKENFNPTTGILVIGNLTNFEQGYYQCTAINRLGNSSCEIDLTSSHPEVG >AI581519_P9 (SEQ IDS NO: 15) residues 23 to 203 (SEQ ID NO: 142)QVTIPDGFVNVTVGSNVTLICIYTTTVASREQLSIQWSFFHKKEMEPISIYFSQGGQAVAIGQFKDRITGSNDPGNASITISHMQPADSGIYICDVNNPPDFLGQNQGILNVSVLVKPSKPLCSVQGRPETGHTISLSCLSALGTPSPVYYWHKLEGRDIVPVKENFTNHRDFGHWKSDKF >AI581519_P10 (SEQ IDS NO: 16) residues 23 To 231 (SEQ ID NO: 143)QVTIPDGFVNVTVGSNVTLICIYTTTVASREQLSIQWSFFHKKEMEPISIYFSQGGQAVAIGQFKDRITGSNDPGNASITISHMQPADSGIYICDVNNPPDFLGQNQGILNVSVLVKPSKPLCSVQGRPETGHTISLSCLSALGTPSPVYYWHKLEGRDIVPVKENFNPTTGILVIGNLTNFEQGYYQCTAINRLGNSSCEIDLTSSRQ, >AI581519_P5 (SEQ IDS NO: 13) residues 26 To 293 (SEQ ID NO: 302)IPDGFVNVTVGSNVTLICIYTTTVASREQLSIQWSFFHKKEMEPISHSSCLSTEGMEEKAVSQCLKMTHARDARGRCSWTSESPWEEGKWPDVEAVKGTLDGQQAELQIYFSQGGQAVAIGQFKDRITGSNDPGNASITISHMQPADSGIYICDVNNPPDFLGQNQGILNVSVLVKPSKPLCSVQGRPETGHTISLSCLSALGTPSPVYYWHKLEGRDIVPVKENFNPTTGILVIGNLTNFEQGYYQCTAINRLGNSSCEIDLT SSHP,

and variants thereof possessing at least 80% sequence identity, morepreferably at least 90% sequence identity therewith and even morepreferably at least 95, 96, 97, 98 or 99% sequence identity therewith.

The term the “soluble ectodomain (ECD)” or “ectodomain” of ILDR1 refersto the polypeptide sequences below or the corresponding nucleic acidsequences (which does not comprise the signal peptide and the TM ofILDR1 protein:

residues 24-105 of AA424839_1_P11 (SEQ ID NO: 24): SEQ ID NO: 296ALSLGQDPSNDCNDNQREVRIVAQRRGQNEPVLGVDYRQRKITIQNRADLVINEVMWWDHGVYYCTIEAPGDTSGDPDKEVK (SEQ ID NO: 296);residues 24-162 of AA424839_P3 (SEQ ID NO: 22) and AA424839_P5 (SEQ ID NO: 21):SEQ ID NO: 75 LLVTVQHTERYVTLFASIILKCDYTTSAQLQDVVVTWRFKSFCKDPIFDYYSASYQAALSLGQDPSNDCNDNQREVRIVAQRRGQNEPVLGVDYRQRKITIQNRADLVINEVMWWDHGVYYCTIEAPGDTSGDPDKEVK:residues 24-457 of AA424839_P7 (SEQ ID NO: 23): SEQ ID NO: 76LLVTVQHTERYVTLFASIILKCDYTTSAQLQDVVVTVVRFKSFCKDPIFDYYSASYQAALSLGQDPSNDCNDNQREVRIVAQRRGQNEPVLGVDYRQRKITIQNPLARHRYMKQAQALGPQMMGKPLYWGADRSSQVSSYPMHPLLQRDLSLPSSLPQMPMTQTTNQPPIANGVLEYLEKELRNLNLAQPLPPDLKGRFGHPCSMLSSLGSEVVERRIIHLPPLIRDLSSSRRTSDSLHQQWLTPIPSRPWDLREGRSHHHYPDFHQELQDRGPKSWALERRELDPSWSGRHRSSRLNGSPIHWSDRDSLSDVPSSSEARWRPSHPPFRSRCQERPRRPSPRESTQRHGRRRRHRSYSPPLPSGLSSWSSEEDKERQPQSWRAHRRGSHSPHWPEEKPPSYRSLDITPGKNSRKKGSVERRSEK DSSHSGRSVVI;residues 50-160 of AA424839_P3 (SEQ ID NO:36): SEQ ID NO: 301AQLQDVVVTWRFKSFCKDPIFDYYSASYQAALSLGQDPSNDCNDNQREVRIVAQRRGQNEPVLGVDYRQRKITIQNRADLVINEVMWWDHGVYYCTIEAP GDTSGDPDKE,

and variants thereof possessing at least 80% sequence identity, morepreferably at least 90% sequence identity therewith and even morepreferably at least 95, 96, 97, 98 or 99% sequence identity therewith.

The term the “soluble ectodomain (ECD)” or “ectodomain” of LOC253012refers to the polypeptide sequences below or the corresponding nucleicacid sequences (which does not comprise the signal peptide and the TM ofLOC253012 protein):

H68654_1_P2 (SEQ ID NO: 35) residues 38-349: (SEQ ID NO: 144)SHTVHGVRGQALYLPVHYGFHTPASDIQIIWLFERPHTMPKYLLGSVNKSVVPDLEYQHKFTMMPPNASLLINPLQFPDEGNYIVKVNIQGNGTLSASQKIQVTVDDPVTKPVVQIHPPSGAVEYVGNMTLTCHVEGGTRLAYQWLKNGRPVHTSSTYSFSPQNNTLHIAPVTKEDIGNYSCLVRNPVSEMESDIIMPIIYYGPYGLQVNSDKGLKVGEVFTVDLGEAILFDCSADSHPPNTYSWIRRTDNTTYIIKHGPRLEVASEKVAQKTMDYVCCAYNNITGRQDETHFTVIITSVGLEKLAQKGKSL;H68654_1_P5, (SEQ ID NO: 36) H68654_1_P7, (SEQ ID NO: 37)H68654_1_P12, (SEQ ID NO: 38) H68654_1_P13, (SEQ ID NO: 39)H68654_1_P14 (SEQ ID NO: 40) residues 19-337: (SEQ ID NO: 145)GLKVTVPSHTVHGVRGQALYLPVHYGFHTPASDIQIIWLFERPHTMPKYLLGSVNKSVVPDLEYQHKFTMMPPNASLLINPLQFPDEGNYIVKVNIQGNGTLSASQKIQVTVDDPVTKPVVQIHPPSGAVEYVGNMTLTCHVEGGTRLAYQWLKNGRPVHTSSTYSFSPQNNTLHIAPVTKEDIGNYSCLVRNPVSEMESDIIMPIIYYGPYGLQVNSDKGLKVGEVFTVDLGEAILFDCSADSHPPNTYSWIRRTDNTTYIIKHGPRLEVASEKVAQKTMDYVCCAYNNITGRQDETHFTVIITSVGLEKLAQKGKSL,H68654_1_P5 (SEQ ID NO: 36) residues 1-335 (SEQ ID NO: 300):MWLKVFTTFLSFATGACSGLKVTVPSHTVHGVRGQALYLPVHYGFHTPASDIQIIWLFERPHTMPKYLLGSVNKSVVPDLEYQHKFTMMPPNASLLINPLQFPDEGNYIVKVNIQGNGTLSASQKIQVTVDDPVTKPVVQIHPPSGAVEYVGNMTLTCHVEGGTRLAYQWLKNGRPVHTSSTYSFSPQNNTLHIAPVTKEDIGNYSCLVRNPVSEMESDIIMPIIYYGPYGLQVNSDKGLKVGEVFTVDLGEAILFDCSADSHPPNTYSWIRRTDNTTYIIKHGPRLEVASEKVAQKTMDYVCCAYNNITGRQDETHFT VIITSVGLEKLAQKGK,

and variants thereof possessing at least 80% sequence identity, morepreferably at least 90% sequence identity therewith and even morepreferably at least 95, 96, 97, 98 or 99% sequence identity therewith.

The term the “soluble ectodomain (ECD)” or “ectodomain” of AI216611refers to the polypeptide sequences below or the corresponding nucleicacid sequences (which does not comprise the signal peptide and the TM ofAI216611 protein):

>AI216611_P0 (SEQ ID NO: 43) From 29 to 147 (SEQ ID NO: 146)LQSQGVSLYIPQATINATVKEDILLSVEYSCHGVPTIEWTYSSNWGTQKIVEWKPGTQANISQSHKDRVCTFDNGSIQLFSVGVRDSGYYVITVTERLGSSQFGTIVLHVSEILYEDLH, >AI216611_P0 (SEQ ID NO: 43) From 1 to 145 (SEQ ID NO: 298)MRPLPSGRRKTRGISLGLFALCLAAARCLQSQGVSLYIPQATINATVKEDILLSVEYSCHGVPTIEWTYSSNWGTQKIVEWKPGTQANISQSHKDRVCTFDNGSIQLFSVGVRDSGYYVITVTERLGSSQFGTIVLHVSEILYED,

and variants thereof possessing at least 80% sequence identity, morepreferably at least 90% sequence identity therewith and even morepreferably at least 95, 96, 97, 98 or 99% sequence identity therewith.

The term the “soluble ectodomain (ECD)” or “ectodomain” of C1ORF32refers to the polypeptide sequences below or the corresponding nucleicacid sequences (which does not comprise the signal peptide and the TM ofC1ORF32 protein):

>H19011_1_P8 (SEQ ID NO: 48) residues 21 to 186 (SEQ ID NO: 147)LQVTVPDKKKVAMLFQPTVLRCHFSTSSHQPAVVQWKFKSYCQDRMGESLGMSSTRAQSLSKRNLEWDPYLDCLDSRRTVRVVASKQGSTVTLGDFYRGREITIVHDADLQIGKLMWGDSGLYYCIITTPDDLEGKNEGSLGLLVLGRTGLLADLLPSFAVEIMPE >H19011_1_P9 (SEQ ID NO: 50) residues 21 to 169 (SEQ ID NO: 148)LQVTVPDKKKVAMLFQPTVLRCHFSTSSHQPAVVQWKFKSYCQDRMGESLGMSSTRAQSLSKRNLEWDPYLDCLDSRRTVRVVASKQGSTVTLGDFYRGREITIVHDADLQIGKLMWGDSGLYYCIITTPDDLEGKNEGSLGLLVLEWV, >H19011_1_P8 (SEQ ID NO: 48) residues 1 to 184 (SEQ ID NO: 299)MDRVLLRWISLFWLTAMVEGLQVTVPDKKKVAMLFQPTVLRCHFSTSSHQPAVVQWKFKSYCQDRMGESLGMSSTRAQSLSKRNLEWDPYLDCLDSRRTVRVVASKQGSTVTLGDFYRGREITIVHDADLQIGKLMWGDSGLYYCIITTPDDLEGKNEDSVELLVLGRTGLLADLLPSFAVEIM,

and variants thereof possessing at least 80% sequence identity, morepreferably at least 90% sequence identity therewith and even morepreferably at least 95, 96, 97, 98 or 99% sequence identity therewith.

The term the “soluble ectodomain (ECD)” or “ectodomain” of FXYD3 refersto the polypeptide sequences below or the corresponding nucleic acidsequences (which does not comprise the signal peptide and the TM ofFXYD3 protein):

>R31375_PO (SEQ ID NO: 70); R31375_P31 (SEQ ID NO: 73)From 21 to 36 (SEQ ID NO: 149)NDLEDKNSPFYYDWHS >R31375_P14 (SEQ ID NO: 72) From 21 to 65 (SEQ ID NO: 150)NDLEDKNSPFYYGAPYIFVKRMGGQMKRTQAGTEVPSTFLLDWHS >R31375_P33 (SEQ ID NO: 74) From 21 to 25 (SEQ ID NO: 151) NDLED, >R31375_P14 (SEQ ID NO: 72) From 1 to 63 (SEQ ID NO: 297)MQKVTLGLLVFLAGFPVLDANDLEDKNSPFYYGAPYIFVKRMGGQMKR TQAGTEVPSTFLLDW,

and variants thereof possessing at least 80% sequence identity, morepreferably at least 90% sequence identity therewith and even morepreferably at least 95, 96, 97, 98 or 99% sequence identity therewith.

The term “immune response” refers to the action of, for example,lymphocytes, antigen presenting cells, phagocytic cells, granulocytes,and soluble macromolecules produced by the above cells or cells producedby the liver or spleen (including antibodies, cytokines, and complement)that results in selective damage to, destruction of, or elimination fromthe human body of invading pathogens, cells or tissues infected withpathogens, cancerous cells, or, in cases of autoimmunity or pathologicalinflammation, normal human cells or tissues.

A “signal, transduction pathway” refers to the biochemical relationshipbetween a variety of signal transduction molecules that play a role inthe transmission of a signal from one portion of a cell to anotherportion of a cell.

As used herein, the phrase “cell surface receptor” includes, forexample, molecules and complexes of molecules capable of receiving asignal and the transmission of such a signal across the plasma membraneof a cell.

The term “antibody” as referred to herein includes whole polyclonal andmonoclonal antibodies and any antigen binding fragment (i.e.,“antigen-binding portion”) or single chains thereof. An “antibody”refers to a glycoprotein comprising at least two heavy (H) chains andtwo light (L) chains inter-connected by disulfide bonds, or an antigenbinding portion thereof. Each heavy chain is comprised of a heavy chainvariable region (abbreviated herein as VH) and a heavy chain constantregion. The heavy chain constant region is comprised of three domains,CH1, CH2 and CH3. Each light chain is comprised of a light chainvariable region (abbreviated herein as VL) and a light chain constantregion. The light chain constant region is comprised of one domain, CL.The VH and VL regions can be further subdivided into regions ofhypervariability, termed complementarity determining regions (CDR),interspersed with regions that are more conserved, termed frameworkregions (FR). Each VH and VL is composed of three CDRs and four FRs,arranged from amino-terminus to carboxy-terminus in the following order:FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions of the heavyand light chains contain a binding domain that interacts with anantigen. The constant regions of the antibodies may mediate the bindingof the immunoglobulin to host tissues or factors, including variouscells of the immune system (e.g., effector cells) and the firstcomponent (Clq) of the classical complement system.

The term “antigen-binding portion” of an antibody (or simply “antibodyportion”), as used herein, refers to one or more fragments of anantibody that retain the ability to specifically bind to an antigen(e.g., VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 proteins orVSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3). It has been shownthat the antigen-binding function of an antibody can be performed byfragments of a full-length antibody. Examples of binding fragmentsencompassed within the term “antigen-binding portion” of an antibodyinclude (i) a Fab fragment, a monovalent fragment consisting of the VLight, V Heavy, Constant light (CL) and CH1 domains; (ii) a F(ab′).2fragment, a bivalent fragment comprising two Fab fragments linked by adisulfide bridge at the hinge region; (iii) a Fd fragment consisting ofthe VH and CH1 domains; (iv) a Fv fragment consisting of the VL and VHdomains of a single arm of an antibody, (v) a dAb fragment (Ward et al.,(1989) Nature 341:544-546), which consists of a VH domain; and (vi) anisolated complementarity determining region (CDR). Furthermore, althoughthe two domains of the Fv fragment, VL and VH, are coded for by separategenes, they can be joined, using recombinant methods, by a syntheticlinker that enables them to be made as a single protein chain in whichthe VL and VH regions pair to form monovalent molecules (known as singlechain Fv (scFv); see e.g., Bird et al. (1988) Science 242:423-426; andHuston et al. (1988) Proc. Natl. Acad. Sci. USA 85:5879-5883). Suchsingle chain antibodies are also intended to be encompassed within theterm “antigen-binding portion” of an antibody. These antibody fragmentsare obtained using conventional techniques known to those with skill inthe art, and the fragments are screened for utility in the same manneras are intact antibodies.

An “isolated antibody”, as used herein, is intended to refer to anantibody that is substantially free of other antibodies having differentantigenic specificities (e.g., an isolated antibody that specificallybinds VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 proteins orVSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3 is substantiallyfree of antibodies that specifically bind antigens other than VSIG1,ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 proteins or VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, or FXYD3, respectively. An isolatedantibody that specifically binds VSIG1, ILDR1, LOC253012, AI216611,C1ORF32, FXYD3 proteins or VSIG1, ILDR1, LOC253012, AI216611, C1ORF32,or FXYD3 may, however, have cross-reactivity to other antigens, such asVSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 proteins or VSIG1,ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3 molecules from otherspecies, respectively. Moreover, an isolated antibody may besubstantially free of other cellular material and/or chemicals.

The terms “monoclonal antibody” or “monoclonal antibody composition” asused herein refer to a preparation of antibody molecules of singlemolecular composition. A monoclonal antibody composition displays asingle binding specificity and affinity for a particular epitope.

The term “human antibody”, as used herein, is intended to includeantibodies having variable regions in which both the framework and CDRregions are derived from human germline immunoglobulin sequences.Furthermore, if the antibody contains a constant region, the constantregion also is derived from human germline immunoglobulin sequences. Thehuman antibodies of the invention may include amino acid residues notencoded by human germline immunoglobulin sequences (e.g., mutationsintroduced by random or site-specific mutagenesis in vitro or by somaticmutation in vivo). However, the term “human antibody”, as used herein,is not intended to include antibodies in which CDR sequences derivedfrom the germline of another mammalian species, such as a mouse, havebeen grafted onto human framework sequences.

The term “human monoclonal antibody” refers to antibodies displaying asingle binding specificity which have variable regions in which both theframework and CDR regions are derived from human germline immunoglobulinsequences. In one embodiment, the human monoclonal antibodies areproduced by a hybridoma which includes a B cell obtained from atransgenic nonhuman animal, e.g., a transgenic mouse, having a genomecomprising a human heavy chain transgene and a light chain transgenefused to an immortalized cell.

The term “recombinant human antibody”, as used herein, includes allhuman antibodies that are prepared, expressed, created or isolated byrecombinant means, such as (a) antibodies isolated from an animal (e.g.,a mouse) that is transgenic or transchromosomal for human immunoglobulingenes or a hybridoma prepared therefrom (described further below), (b)antibodies isolated from a host cell transformed to express the humanantibody, e.g., from a transfectoma, (c) antibodies isolated from arecombinant, combinatorial human antibody library, and (d) antibodiesprepared, expressed, created or isolated by any other means that involvesplicing of human immunoglobulin gene sequences to other DNA sequences.Such recombinant human antibodies have variable regions in which theframework and CDR regions are derived from human germline immunoglobulinsequences. In certain embodiments, however, such recombinant humanantibodies can be subjected to in vitro mutagenesis (or, when an animaltransgenic for human Ig sequences is used, in vivo somatic mutagenesis)and thus the amino acid sequences of the VH and VL regions of therecombinant antibodies are sequences that, while derived from andrelated to human germline VH and VL sequences, may not naturally existwithin the human antibody germline repertoire in vivo.

As used herein, “isotype” refers to the antibody class (e.g., IgM orIgG1) that is encoded by the heavy chain constant region genes.

The phrases “an antibody recognizing an antigen” and “an antibodyspecific for an antigen” are used interchangeably herein with the term“an antibody which binds specifically to an antigen.”

As used herein, an antibody that “specifically binds to human VSIG1,ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 proteins or VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, or FXYD3 is intended to refer to anantibody that binds to human VSIG1, ILDR1, LOC253012, AI216611, C1ORF32,FXYD3 proteins or VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3,respectively, preferably one with a KD of 5×10−8 M or less, morepreferably 3×10−8 M or less, and even more preferably 1×.10−9 M or less.

The term “K-assoc” or “Ka”, as used herein, is intended to refer to theassociation rate of a particular antibody-antigen interaction, whereasthe term “Kdiss” or “Kd,” as used herein, is intended to refer to thedissociation rate of a particular antibody-antigen interaction. The term“KD”, as used herein, is intended to refer to the dissociation constant,which is obtained from the ratio of Kd to Ka (i.e., Kd/Ka) and isexpressed as a molar concentration (M). KD values for antibodies can bedetermined using methods well established in the art. A preferred methodfor determining the KD of an antibody is by using surface Plasmonresonance, preferably using a biosensor system such as a Biacore®system.

As used herein, the term “high affinity” for an IgG antibody refers toan antibody having a KD of 10−8 M or less, more preferably 10−9 M orless and even more preferably 10−10 M or less for a target antigen.However, “high affinity” binding can vary for other antibody isotypes.For example, “high affinity” binding for an IgM isotype refers to anantibody having a KD of 10−7 M or less, more preferably 10−8 M or less.

As used herein, the term “subject” includes any human or nonhumananimal. The term “nonhuman animal” includes all vertebrates, e.g.,mammals and non-mammals, such as nonhuman primates, sheep, dogs, cats,horses, cows chickens, amphibians, reptiles, etc.

As used herein, the term “tail” refers to a peptide sequence at the endof an amino acid sequence that is unique to a splice variant accordingto the present invention. Therefore, a splice variant having such a tailmay optionally be considered as a chimera, in that at least a firstportion of the splice variant is typically highly homologous (often 100%identical) to a portion of the corresponding known protein, while atleast a second portion of the variant comprises the tail.

As used herein, the term “head” refers to a peptide sequence at thebeginning of an amino acid sequence that is unique to a splice variantaccording to the present invention. Therefore, a splice variant havingsuch a head may optionally be considered as a chimera, in that at leasta first portion of the splice variant comprises the head, while at leasta second portion is typically highly homologous (often 100% identical)to a portion of the corresponding known protein.

As used herein, the term “an edge portion” refers to a connectionbetween two portions of a splice variant according to the presentinvention that were not joined in the wild type or known protein. Anedge may optionally arise due to a join between the above “knownprotein” portion of a variant and the tail, for example, and/or mayoccur if an internal portion of the wild type sequence is no longerpresent, such that two portions of the sequence are now contiguous inthe splice variant that were not contiguous in the known protein. A“bridge” may optionally be an edge portion as described above, but mayalso include a join between a head and a “known protein” portion of avariant, or a join between a tail and a “known protein” portion of avariant, or a join between an insertion and a “known protein” portion ofa variant.

In some embodiments, a bridge between a tail or a head or a uniqueinsertion, and a “known protein” portion of a variant, comprises atleast about 10 amino acids, or in some embodiments at least about 20amino acids, or in some embodiments at least about 30 amino acids, or insome embodiments at least about 40 amino acids, in which at least oneamino acid is from the tail/head/insertion and at least one amino acidis from the “known protein” portion of a variant. In some embodiments,the bridge may comprise any number of amino acids from about 10 to about40 amino acids (for example, 10, 11, 12, 13 . . . 37, 38, 39, 40 aminoacids in length, or any number in between).

It should be noted that a bridge cannot be extended beyond the length ofthe sequence in either direction, and it should be assumed that everybridge description is to be read in such manner that the bridge lengthdoes not extend beyond the sequence itself.

Furthermore, bridges are described with regard to a sliding window incertain contexts below. For example, certain descriptions of the bridgesfeature the following format: a bridge between two edges (in which aportion of the known protein is not present in the variant) mayoptionally be described as follows: a bridge portion of CONTIG-NAME_P1(representing the name of the protein), comprising a polypeptide havinga length “n”, wherein n is at least about 10 amino acids in length,optionally at least about 20 amino acids in length, preferably at leastabout 30 amino acids in length, more preferably at least about 40 aminoacids in length and most preferably at least about 50 amino acids inlength, wherein at least two amino acids comprise XX (2 amino acids inthe center of the bridge, one from each end of the edge), having astructure as follows (numbering according to the sequence ofCONTIG-NAME_P1): a sequence starting from any of amino acid numbers 49−xto 49 (for example); and ending at any of amino acid numbers50+((n−2)−x) (for example), in which x varies from 0 to n−2. In thisexample, it should also be read as including bridges in which n is anynumber of amino acids between 10-50 amino acids in length. Furthermore,the bridge polypeptide cannot extend beyond the sequence, so it shouldbe read such that 49−x (for example) is not less than 1, nor50+((n−2)−x) (for example) greater than the total sequence length.

Various aspects of the invention are described in further detail in thefollowing subsections.

Nucleic Acids

A “nucleic acid fragment” or an “oligonucleotide” or a “polynucleotide”are used herein interchangeably to refer to a polymer of nucleic acidresidues. A polynucleotide sequence of the present invention refers to asingle or double stranded nucleic acid sequences which is isolated andprovided in the form of an RNA sequence, a complementary polynucleotidesequence (cDNA), a genomic polynucleotide sequence and/or a compositepolynucleotide sequences (e.g., a combination of the above).

Thus, the present invention encompasses nucleic acid sequences describedhereinabove; fragments thereof, sequences hybridizable therewith,sequences homologous thereto [e.g., at least 90%, at least 95, 96, 97,98 or 99% or more identical to the nucleic acid sequences set forthherein], sequences encoding similar polypeptides with different codonusage, altered sequences characterized by mutations, such as deletion,insertion or substitution of one or more nucleotides, either naturallyoccurring or man induced, either randomly or in a targeted fashion. Thepresent invention also encompasses homologous nucleic acid sequences(i.e., which form a part of a polynucleotide sequence of the presentinvention), which include sequence regions unique to the polynucleotidesof the present invention.

In cases where the polynucleotide sequences of the present inventionencode previously unidentified polypeptides, the present invention alsoencompasses novel polypeptides or portions thereof, which are encoded bythe isolated polynucleotide and respective nucleic acid fragmentsthereof described hereinabove.

Thus, the present invention also encompasses polypeptides encoded by thepolynucleotide sequences of the present invention. The present inventionalso encompasses homologues of these polypeptides, such homologues canbe at least 90%, at least 95, 96, 97, 98 or 99% or more homologous tothe amino acid sequences set forth below, as can be determined usingBlastP software of the National Center of Biotechnology Information(NCBI) using default parameters. Finally, the present invention alsoencompasses fragments of the above described polypeptides andpolypeptides having mutations, such as deletions, insertions orsubstitutions of one or more amino acids, either naturally occurring orman induced, either randomly or in a targeted fashion.

As mentioned hereinabove, biomolecular sequences of the presentinvention can be efficiently utilized as tissue or pathological markersand as putative drugs or drug targets for treating or preventing adisease.

Oligonucleotides designed for carrying out the methods of the presentinvention for any of the sequences provided herein (designed asdescribed above) can be generated according to any oligonucleotidesynthesis method known in the art such as enzymatic synthesis or solidphase synthesis. Equipment and reagents for executing solid-phasesynthesis are commercially available from, for example, AppliedBiosystems. Any other means for such synthesis may also be employed; theactual synthesis of the oligonucleotides is well within the capabilitiesof one skilled in the art.

Oligonucleotides used according to this aspect of the present inventionare those having a length selected from a range of about 10 to about 200bases preferably about 15 to about 150 bases, more preferably about 20to about 100 bases, most preferably about 20 to about 50 bases.

The oligonucleotides of the present invention may comprise heterocyclicnucleosides consisting of purines and the pyrimidines bases, bonded in a3′ to 5′ phosphodiester linkage.

Preferable oligonucleotides are those modified in either backbone,internucleoside linkages or bases, as is broadly described hereinunder.Such modifications can oftentimes facilitate oligonucleotide uptake andresistivity to intracellular conditions.

Specific examples of preferred oligonucleotides useful according to thisaspect of the present invention include oligonucleotides containingmodified backbones or non-natural internucleoside linkages.Oligonucleotides having modified backbones include those that retain aphosphorus atom in the backbone, as disclosed in U.S. Pat. Nos.4,469,863; 4,476,301; 5,023,243; 5,177,196; 5,188,897; 5,264,423;5,276,019; 5,278,302; 5,286,717; 5,321,131; 5,399,676; 5,405,939;5,453,496; 5,455,233; 5,466,677; 5,476,925; 5,519,126; 5,536,821;5,541,306; 5,550,111; 5,563,253; 5,571,799; 5,587,361; and 5,625,050.

Preferred modified oligonucleotide backbones include, for example,phosphorothioates, chiral phosphorothioates, phosphorodithioates,phosphotriesters, aminoalkyl phosphotriesters, methyl and other alkylphosphonates including 3′-alkylene phosphonates and chiral phosphonates,phosphinates, phosphoramidates including 3′-amino phosphoramidate andaminoalkylphosphoramidates, thionophosphoramidates,thionoalkylphosphonates, thionoalkylphosphotriesters, andboranophosphates having normal 3′-5′ linkages, 2′-5′ linked analogs ofthese, and those having inverted polarity wherein the adjacent pairs ofnucleoside units are linked 3′-5′ to 5′-3′ or 2′-5′ to 5′-2′. Varioussalts, mixed salts and free acid forms can also be used.

Alternatively, modified oligonucleotide backbones that do not include aphosphorus atom therein have backbones that are formed by short chainalkyl or cycloalkyl internucleoside linkages, mixed heteroatom and alkylor cycloalkyl internucleoside linkages, or one or more short chainheteroatomic or heterocyclic internucleoside linkages. These includethose having morpholino linkages (formed in part from the sugar portionof a nucleoside); siloxane backbones; sulfide, sulfoxide and sulfonebackbones; formacetyl and thioformacetyl backbones; methylene formacetyland thioformacetyl backbones; alkene containing backbones; sulfamatebackbones; methyleneimino and methylenehydrazino backbones; sulfonateand sulfonamide backbones; amide backbones; and others having mixed N,O, S and CH2 component parts, as disclosed in U.S. Pat. Nos. 5,034,506;5,166,315; 5,185,444; 5,214,134; 5,216,141; 5,235,033; 5,264,562;5,264,564; 5,405,938; 5,434,257; 5,466,677; 5,470,967; 5,489,677;5,541,307; 5,561,225; 5,596,086; 5,602,240; 5,610,289; 5,602,240;5,608,046; 5,610,289; 5,618,704; 5,623,070; 5,663,312; 5,633,360;5,677,437; and 5,677,439.

Other oligonucleotides which can be used according to the presentinvention, are those modified in both sugar and the internucleosidelinkage, i.e., the backbone, of the nucleotide units are replaced withnovel groups. The base units are maintained for complementation with theappropriate polynucleotide target. An example for such anoligonucleotide mimetic, includes peptide nucleic acid (PNA). A PNAoligonucleotide refers to an oligonucleotide where the sugar-backbone isreplaced with an amide containing backbone, in particular anaminoethylglycine backbone. The bases are retained and are bounddirectly or indirectly to aza nitrogen atoms of the amide portion of thebackbone. United States patents that teach the preparation of PNAcompounds include, but are not limited to, U.S. Pat. Nos. 5,539,082;5,714,331; and 5,719,262, each of which is herein incorporated byreference. Other backbone modifications, which can be used in thepresent invention are disclosed in U.S. Pat. No. 6,303,374.

Oligonucleotides of the present invention may also include basemodifications or substitutions. As used herein, “unmodified” or“natural” bases include the purine bases adenine (A) and guanine (G),and the pyrimidine bases thymine (T), cytosine (C) and uracil (U).Modified bases include but are not limited to other synthetic andnatural bases such as 5-methylcytosine (5-me-C), 5-hydroxymethylcytosine, xanthine, hypoxanthine, 2-aminoadenine, 6-methyl and otheralkyl derivatives of adenine and guanine, 2-propyl and other alkylderivatives of adenine and guanine, 2-thiouracil, 2-thiothymine and2-thiocytosine, 5-halouracil and cytosine, 5-propynyl uracil andcytosine, 6-azo uracil, cytosine and thymine, 5-uracil (pseudouracil),4-thiouracil, 8-halo, 8-amino, 8-thiol, 8-thioalkyl, 8-hydroxyl andother 8-substituted adenines and guanines, 5-halo particularly 5-bromo,5-trifluoromethyl and other 5-substituted uracils and cytosines,7-methylguanine and 7-methyladenine, 8-azaguanine and 8-azaadenine,7-deazaguanine and 7-deazaadenine and 3-deazaguanine and 3-deazaadenine.Further bases include those disclosed in U.S. Pat. No. 3,687,808, thosedisclosed in The Concise Encyclopedia Of Polymer Science andEngineering, pages 858-859, Kroschwitz, J. I., ed. John Wiley & Sons,1990, those disclosed by Englisch et al., Angewandte Chemie,International Edition, 1991, 30, 613, and those disclosed by Sanghvi, Y.S., Chapter 15, Antisense Research and Applications, pages 289-302,Crooke, S. T. and Lebleu, B., ed., CRC Press, 1993. Such bases areparticularly useful for increasing the binding affinity of theoligomeric compounds of the invention. These include 5-substitutedpyrimidines, 6-azapyrimidines and N-2, N-6 and O-6 substituted purines,including 2-aminopropyladenine, 5-propynyluracil and 5-propynylcytosine.5-methylcytosine substitutions have been shown to increase nucleic acidduplex stability by 0.6-1.2° C. [Sanghvi Y S et al. (1993) AntisenseResearch and Applications, CRC Press, Boca Raton 276-278] and arepresently preferred base substitutions, even more particularly whencombined with 2′-O-methoxyethyl sugar modifications.

Another modification of the oligonucleotides of the invention involveschemically linking to the oligonucleotide one or more moieties orconjugates, which enhance the activity, cellular distribution orcellular uptake of the oligonucleotide. Such moieties include but arenot limited to lipid moieties such as a cholesterol moiety, cholic acid,a thioether, e.g., hexyl-5-tritylthiol, a thiocholesterol, an aliphaticchain, e.g., dodecandiol or undecyl residues, a phospholipid, e.g.,di-hexadecyl-rac-glycerol or triethylammonium1,2-di-O-hexadecyl-rac-glycero-3-H-phosphonate, a polyamine or apolyethylene glycol chain, or adamantane acetic acid, a palmityl moiety,or an octadecylamine or hexylamino-carbonyl-oxycholesterol moiety, asdisclosed in U.S. Pat. No. 6,303,374.

It is not necessary for all positions in a given oligonucleotidemolecule to be uniformly modified, and in fact more than one of theaforementioned modifications may be incorporated in a single compound oreven at a single nucleoside within an oligonucleotide.

Peptides

The terms “polypeptide,” “peptide” and “protein” are usedinterchangeably herein to refer to a polymer of amino acid residues. Theterms apply to amino acid polymers in which one or more amino acidresidue is an analog or mimetic of a corresponding naturally occurringamino acid, as well as to naturally occurring amino acid polymers.Polypeptides can be modified, e.g., by the addition of carbohydrateresidues to form glycoproteins. The terms “polypeptide,” “peptide” and“protein” include glycoproteins, as well as non-glycoproteins.

Polypeptide products can be biochemically synthesized such as byemploying standard solid phase techniques. Such methods includeexclusive solid phase synthesis, partial solid phase synthesis methods,fragment condensation, classical solution synthesis. These methods arepreferably used when the peptide is relatively short (i.e., 10 kDa)and/or when it cannot be produced by recombinant techniques (i.e., notencoded by a nucleic acid sequence) and therefore involves differentchemistry.

Solid phase polypeptide synthesis procedures are well known in the artand further described by John Morrow Stewart and Janis Dillaha Young,Solid Phase Peptide Syntheses (2nd Ed., Pierce Chemical Company, 1984).

Synthetic polypeptides can be purified by preparative high performanceliquid chromatography [Creighton T. (1983) Proteins, structures andmolecular principles. WH Freeman and Co. N.Y.] and the composition ofwhich can be confirmed via amino acid sequencing.

In cases where large amounts of a polypeptide are desired, it can begenerated using recombinant techniques such as described by Bitter etal., (1987) Methods in Enzymol. 153:516-544, Studier et al. (1990)Methods in Enzymol. 185:60-89, Brisson et al. (1984) Nature 310:511-514,Takamatsu et al. (1987) EMBO J. 6:307-311, Coruzzi et al. (1984) EMBO J.3:1671-1680 and Brogli et al., (1984) Science 224:838-843, Gurley et al.(1986) Mol. Cell. Biol. 6:559-565 and Weissbach & Weissbach, 1988,Methods for Plant Molecular Biology, Academic Press, NY, Section VIII,pp 421-463.

It will be appreciated that peptides identified according to theteachings of the present invention may be degradation products,synthetic peptides or recombinant peptides as well as peptidomimetics,typically, synthetic peptides and peptoids and semipeptoids which arepeptide analogs, which may have, for example, modifications renderingthe peptides more stable while in a body or more capable of penetratinginto cells. Such modifications include, but are not limited to Nterminus modification, C terminus modification, peptide bondmodification, including, but not limited to, CH2-NH, CH2-S, CH2-S═O,O═C—NH, CH2-O, CH2-CH2, S═C—NH, CH═CH or CF═CH, backbone modifications,and residue modification. Methods for preparing peptidomimetic compoundsare well known in the art and are specified, for example, inQuantitative Drug Design, C.A. Ramsden Gd., Chapter 17.2, F. ChoplinPergamon Press (1992), which is incorporated by reference as if fullyset forth herein. Further details in this respect are providedhereinunder.

Peptide bonds (—CO—NH—) within the peptide may be substituted, forexample, by N-methylated bonds (—N(CH3)-CO—), ester bonds(—C(R)H—C—O—O—C(R)—N—), ketomethylen bonds (—CO—CH2-), α-aza bonds(—NH—N(R)—CO—), wherein R is any alkyl, e.g., methyl, carba bonds(—CH2-NH—), hydroxyethylene bonds (—CH(OH)—CH₂-), thioamide bonds(—CS—NH—), olefinic double bonds (—CH═CH—), retro amide bonds (—NH—CO—),peptide derivatives (—N(R)—CH2-CO—), wherein R is the “normal” sidechain, naturally presented on the carbon atom.

These modifications can occur at any of the bonds along the peptidechain and even at several (2-3) at the same time.

Natural aromatic amino acids, Trp, Tyr and Phe, may be substituted bysynthetic non-natural acid such as Phenylglycine, TIC, naphthylelanine(Nol), ring-methylated derivatives of Phe, halogenated derivatives ofPhe or o-methyl-Tyr.

In addition to the above, the peptides of the present invention may alsoinclude one or more modified amino acids or one or more non-amino acidmonomers (e.g. fatty acids, complex carbohydrates etc).

As used herein in the specification and in the claims section below theterm “amino acid” or “amino acids” is understood to include the 20naturally occurring amino acids; those amino acids often modifiedpost-translationally in vivo, including, for example, hydroxyproline,phosphoserine and phosphothreonine; and other unusual amino acidsincluding, but not limited to, 2-aminoadipic acid, hydroxylysine,isodesmosine, nor-valine, nor-leucine and ornithine. Furthermore, theterm “amino acid” includes both D- and L-amino acids.

Since the peptides of the present invention are preferably utilized intherapeutics which require the peptides to be in soluble form, thepeptides of the present invention preferably include one or morenon-natural or natural polar amino acids, including but not limited toserine and threonine which are capable of increasing peptide solubilitydue to their hydroxyl-containing side chain.

The peptides of the present invention are preferably utilized in alinear form, although it will be appreciated that in cases wherecyclization does not severely interfere with peptide characteristics,cyclic forms of the peptide can also be utilized.

The peptides of the present invention can be biochemically synthesizedsuch as by using standard solid phase techniques. These methods includeexclusive solid phase synthesis, partial solid phase synthesis methods,fragment condensation, classical solution synthesis. These methods arepreferably used when the peptide is relatively short (i.e., 10 kDa)and/or when it cannot be produced by recombinant techniques (i.e., notencoded by a nucleic acid sequence) and therefore involves differentchemistry.

Solid phase peptide synthesis procedures are well known in the art andfurther described by John Morrow Stewart and Janis Dillaha Young, SolidPhase Peptide Syntheses (2nd Ed., Pierce Chemical Company, 1984).

Synthetic peptides can be purified by preparative high performanceliquid chromatography [Creighton T. (1983) Proteins, structures andmolecular principles. WH Freeman and Co. N.Y.] and the composition ofwhich can be confirmed via amino acid sequencing.

In cases where large amounts of the peptides of the present inventionare desired, the peptides of the present invention can be generatedusing recombinant techniques such as described by Bitter et al., (1987)Methods in Enzymol. 153:516-544, Studier et al. (1990) Methods inEnzymol. 185:60-89, Brisson et al. (1984) Nature 310:511-514, Takamatsuet al. (1987) EMBO J. 6:307-311, Coruzzi et al. (1984) EMBO J.3:1671-1680 and Brogli et al., (1984) Science 224:838-843, Gurley et al.(1986) Mol. Cell. Biol. 6:559-565 and Weissbach & Weissbach, 1988,Methods for Plant Molecular Biology, Academic Press, NY, Section VIII,pp 421-463.

Expression Systems

To enable cellular expression of the polynucleotides of the presentinvention, a nucleic acid construct according to the present inventionmay be used, which includes at least a coding region of one of the abovenucleic acid sequences, and further includes at least one cis actingregulatory element. As used herein, the phrase “cis acting regulatoryelement” refers to a polynucleotide sequence, preferably a promoter,which binds a trans acting regulator and regulates the transcription ofa coding sequence located downstream thereto.

Any suitable promoter sequence can be used by the nucleic acid constructof the present invention.

Preferably, the promoter utilized by the nucleic acid construct of thepresent invention is active in the specific cell population transformed.Examples of cell type-specific and/or tissue-specific promoters includepromoters such as albumin that is liver specific [Pinkert et al., (1987)Genes Dev. 1:268-277], lymphoid specific promoters [Calame et al.,(1988) Adv. Immunol. 43:235-275]; in particular promoters of T-cellreceptors [Winoto et al., (1989) EMBO J. 8:729-733] and immunoglobulins;[Banerji et al. (1983) Cell 33729-740], neuron-specific promoters suchas the neurofilament promoter [Byrne et al. (1989) Proc. Natl. Acad.Sci. USA 86:5473-5477], pancreas-specific promoters [Edlunch et al.(1985) Science 230:912-916] or mammary gland-specific promoters such asthe milk whey promoter (U.S. Pat. No. 4,873,316 and European ApplicationPublication No. 264,166). The nucleic acid construct of the presentinvention can further include an enhancer, which can be adjacent ordistant to the promoter sequence and can function in up regulating thetranscription therefrom.

The nucleic acid construct of the present invention preferably furtherincludes an appropriate selectable marker and/or an origin ofreplication. Preferably, the nucleic acid construct utilized is ashuttle vector, which can propagate both in E. coli (wherein theconstruct comprises an appropriate selectable marker and origin ofreplication) and be compatible for propagation in cells, or integrationin a gene and a tissue of choice. The construct according to the presentinvention can be, for example, a plasmid, a bacmid, a phagemid, acosmid, a phage, a virus or an artificial chromosome.

Examples of suitable constructs include, but are not limited to, pcDNA3,pcDNA3.1 (+/−), pGL3, PzeoSV2 (+/−), pDisplay, pEF/myc/cyto,pCMV/myc/cyto each of which is commercially available from InvitrogenCo. (www.invitrogen.com). Examples of retroviral vector and packagingsystems are those sold by Clontech, San Diego, Calif., including Retro-Xvectors pLNCX and pLXSN, which permit cloning into multiple cloningsites and the transgene is transcribed from CMV promoter. Vectorsderived from Mo-MuLV are also included such as pBabe, where thetransgene will be transcribed from the 5′LTR promoter.

Currently preferred in vivo nucleic acid transfer techniques includetransfection with viral or non-viral constructs, such as adenovirus,lentivirus, Herpes simplex I virus, or adeno-associated virus (AAV) andlipid-based systems. Useful lipids for lipid-mediated transfer of thegene are, for example, DOTMA, DOPE, and DC-Chol [Tonkinson et al.,Cancer Investigation, 14(1): 54-65 (1996)]. The most preferredconstructs for use in gene therapy are viruses, most preferablyadenoviruses, AAV, lentiviruses, or retroviruses. A viral construct suchas a retroviral construct includes at least one transcriptionalpromoter/enhancer or locus-defining elements, or other elements thatcontrol gene expression by other means such as alternate splicing,nuclear RNA export, or post-translational modification of messenger.Such vector constructs also include a packaging signal, long terminalrepeats (LTRs) or portions thereof, and positive and negative strandprimer binding sites appropriate to the virus used, unless it is alreadypresent in the viral construct. In addition, such a construct typicallyincludes a signal sequence for secretion of the peptide from a host cellin which it is placed. Preferably the signal sequence for this purposeis a mammalian signal sequence or the signal sequence of thepolypeptides of the present invention. Optionally, the construct mayalso include a signal that directs polyadenylation, as well as one ormore restriction sites and a translation termination sequence. By way ofexample, such constructs will typically include a 5′ LTR, a tRNA bindingsite, a packaging signal, an origin of second-strand DNA synthesis, anda 3′ LTR or a portion thereof. Other vectors can be used that arenon-viral, such as cationic lipids, polylysine, and dendrimers.

Recombinant Expression Vectors and Host Cells

Another aspect of the invention pertains to vectors, preferablyexpression vectors, containing a nucleic acid encoding a protein of theinvention, or derivatives, fragments, analogs or homologs thereof. Asused herein, the term “vector” refers to a nucleic acid molecule capableof transporting another nucleic acid to which it has been linked. Onetype of vector is a “plasmid”, which refers to a circular doublestranded DNA loop into which additional DNA segments can be ligated.Another type of vector is a viral vector, wherein additional DNAsegments can be ligated into the viral genome. Certain vectors arecapable of autonomous replication in a host cell into which they areintroduced (e.g., bacterial vectors having a bacterial origin ofreplication and episomal mammalian vectors). Other vectors (e.g.,non-episomal mammalian vectors) are integrated into the genome of a hostcell upon introduction into the host cell, and thereby are replicatedalong with the host genome. Moreover, certain vectors are capable ofdirecting the expression of genes to which they are operatively-linked.Such vectors are referred to herein as “expression vectors”. In general,expression vectors of utility in recombinant DNA techniques are often inthe form of plasmids. In the present specification, “plasmid” and“vector” can be used interchangeably as the plasmid is the most commonlyused form of vector. However, the invention is intended to include suchother forms of expression vectors, such as viral vectors (e.g.,replication defective retroviruses, adenoviruses and adeno-associatedviruses), which serve equivalent functions.

The recombinant expression vectors of the invention comprise a nucleicacid of the invention in a form suitable for expression of the nucleicacid in a host cell, which means that the recombinant expression vectorsinclude one or more regulatory sequences, selected on the basis of thehost cells to be used for expression, that is operatively-linked to thenucleic acid sequence to be expressed. Within a recombinant expressionvector, “operably-linked” is intended to mean that the nucleotidesequence of interest is linked to the regulatory sequences in a mannerthat allows for expression of the nucleotide sequence (e.g., in an invitro transcription/translation system or in a host cell when the vectoris introduced into the host cell).

The term “regulatory sequence” is intended to includes promoters,enhancers and other expression control elements (e.g., polyadenylationsignals). Such regulatory sequences are described, for example, inGoeddel, Gene Expression Technology Methods in Enzymology 185, AcademicPress, San Diego, Calif. (1990). Regulatory sequences include those thatdirect constitutive expression of a nucleotide sequence in many types ofhost cell and those that direct expression of the nucleotide sequenceonly in certain host cells (e.g., tissue-specific regulatory sequences).It will be appreciated by those skilled in the art that the design ofthe expression vector can depend on such factors as the choice of thehost cell to be transformed, the level of expression of protein desired,etc. The expression vectors of the invention can be introduced into hostcells to thereby produce proteins or peptides, including fusion proteinsor peptides, encoded by nucleic acids as described herein.

The recombinant expression vectors of the invention can be designed forproduction of variant proteins in prokaryotic or eukaryotic cells. Forexample, proteins of the invention can be expressed in bacterial cellssuch as Escherichia coli, insect cells (using baculovirus expressionvectors) yeast cells or mammalian cells. Suitable host cells arediscussed further in Goeddel, Gene Expression Technology: Methods inEnzymology 185, Academic Press, San Diego, Calif. (1990). Alternatively,the recombinant expression vector can be transcribed and translated invitro, for example using T7 promoter regulatory sequences and T7polymerase.

Expression of proteins in prokaryotes is most often carried out inEscherichia coli with vectors containing constitutive or induciblepromoters directing the expression of either fusion or non-fusionproteins. Fusion vectors add a number of amino acids to a proteinencoded therein, to the amino or C terminus of the recombinant protein.Such fusion vectors typically serve three purposes: (i) to increaseexpression of recombinant protein; (ii) to increase the solubility ofthe recombinant protein; and (iii) to aid in the purification of therecombinant protein by acting as a ligand in affinity purification.Often, in fusion expression vectors, a proteolytic cleavage site isintroduced at the junction of the fusion moiety and the recombinantprotein to enable separation of the recombinant protein from the fusionmoiety subsequent to purification of the fusion protein. Such enzymes,and their cognate recognition sequences, include Factor Xa, thrombin,PreScission, TEV and enterokinase. Typical fusion expression vectorsinclude pGEX (Pharmacia Biotech Inc; Smith and Johnson, 1988. Gene 67:31-40), pMAL (New England Biolabs, Beverly, Mass.) and pRIT5 (Pharmacia,Piscataway, N.J.) that fuse glutathione S-transferase (GST), maltose Ebinding protein, or protein A, respectively, to the target recombinantprotein.

Examples of suitable inducible non-fusion E. coli expression vectorsinclude pTrc (Amrann et al., (1988) Gene 69:301-315) and pET 11d(Studier et al., Gene Expression Technology: Methods in Enzymology 185,Academic Press, San Diego, Calif. (1990) 60-89)—not accurate, pET11a-dhave N terminal T7 tag.

One strategy to maximize recombinant protein expression in E. coli is toexpress the protein in a host bacterium with an impaired capacity toproteolytically cleave the recombinant protein. See, e.g., Gottesman,Gene Expression Technology: Methods in Enzymology 185, Academic Press,San Diego, Calif. (1990) 119-128. Another strategy is to alter thenucleic acid sequence of the nucleic acid to be inserted into anexpression vector so that the individual codons for each amino acid arethose preferentially utilized in E. coli (see, e.g., Wada, et al., 1992.Nucl. Acids Res. 20: 2111-2118). Such alteration of nucleic acidsequences of the invention can be carried out by standard DNA synthesistechniques. Another strategy to solve codon bias is by using BL21-codonplus bacterial strains (Invitrogen) or Rosetta bacterial strain(Novagen), these strains contain extra copies of rare E. coli tRNAgenes.

In another embodiment, the expression vector encoding for the protein ofthe invention is a yeast expression vector. Examples of vectors forexpression in yeast Saccharomyces cerevisiae include pYepSec1 (Baldari,et al., 1987. EMBO J. 6: 229-234), pMFa (Kurjan and Herskowitz, 1982.Cell 30: 933-943), pJRY88 (Schultz et al., 1987. Gene 54: 113-123),pYES2 (Invitrogen Corporation, San Diego, Calif.), and picZ (InVitrogenCorp, San Diego, Calif.).

Alternatively, polypeptides of the present invention can be produced ininsect cells using baculovirus expression vectors. Baculovirus vectorsavailable for expression of proteins in cultured insect cells (e.g., SF9cells) include the pAc series (Smith, et al., 1983. Mol. Cell. Biol. 3:2156-2165) and the pVL series (Lucklow and Summers, 1989. Virology 170:31-39).

In yet another embodiment, a nucleic acid of the invention is expressedin mammalian cells using a mammalian expression vector. Examples ofmammalian expression vectors include pCDM8 (Seed, 1987. Nature 329: 840)and pMT2PC (Kaufman, et al., 1987. EMBO J. 6: 187-195), pIRESpuro(Clontech), pUB6 (Invitrogen), pCEP4 (Invitrogen) pREP4 (Invitrogen),pcDNA3 (Invitrogen). When used in mammalian cells, the expressionvector's control functions are often provided by viral regulatoryelements. For example, commonly used promoters are derived from polyoma,adenovirus 2, cytomegalovirus, Rous Sarcoma Virus, and simian virus 40.For other suitable expression systems for both prokaryotic andeukaryotic cells see, e.g., Chapters 16 and 17 of Sambrook, et al.,Molecular Cloning: A Laboratory Manual. 2nd ed., Cold Spring HarborLaboratory, Cold Spring Harbor Laboratory Press, Cold Spring Harbor,N.Y., 1989.

In another embodiment, the recombinant mammalian expression vector iscapable of directing expression of the nucleic acid preferentially in aparticular cell type (e.g., tissue-specific regulatory elements are usedto express the nucleic acid). Tissue-specific regulatory elements areknown in the art. Non-limiting examples of suitable tissue-specificpromoters include the albumin promoter (liver-specific; Pinkert, et al.,1987. Genes Dev. 1: 268-277), lymphoid-specific promoters (Calame andEaton, 1988. Adv. Immunol. 43: 235-275), in particular promoters of Tcell receptors (Winoto and Baltimore, 1989. EMBO J. 8: 729-733) andimmunoglobulins (Banerji, et al., 1983. Cell 33: 729-740; Queen andBaltimore, 1983. Cell 33: 741-748), neuron-specific promoters (e.g., theneurofilament promoter; Byrne and Ruddle, 1989. Proc. Natl. Acad. Sci.USA 86: 5473-5477), pancreas-specific promoters (Edlund, et al., 1985.Science 230: 912-916), and mammary gland-specific promoters (e.g., milkwhey promoter; U.S. Pat. No. 4,873,316 and European ApplicationPublication No. 264,166). Developmentally-regulated promoters are alsoencompassed, e.g., the murine hox promoters (Kessel and Gruss, 1990.Science 249: 374-379) and the alpha-fetoprotein promoter (Campes andTilghman, 1989. Genes Dev. 3: 537-546).

The invention further provides a recombinant expression vectorcomprising a DNA molecule of the invention cloned into the expressionvector in an antisense orientation. That is, the DNA molecule isoperatively-linked to a regulatory sequence in a manner that allows forexpression (by transcription of the DNA molecule) of an RNA moleculethat is antisense to mRNA encoding for protein of the invention.Regulatory sequences operatively linked to a nucleic acid cloned in theantisense orientation can be chosen that direct the continuousexpression of the antisense RNA molecule in a variety of cell types, forinstance viral promoters and/or enhancers, or regulatory sequences canbe chosen that direct constitutive, tissue specific or cell typespecific expression of antisense RNA. The antisense expression vectorcan be in the form of a recombinant plasmid, phagemid or attenuatedvirus in which antisense nucleic acids are produced under the control ofa high efficiency regulatory region, the activity of which can bedetermined by the cell type into which the vector is introduced. For adiscussion of the regulation of gene expression using antisense genessee, e.g., Weintraub, et al., “Antisense RNA as a molecular tool forgenetic analysis,” Reviews-Trends in Genetics, Vol. 1(1) 1986.

Another aspect of the invention pertains to host cells into which arecombinant expression vector of the invention has been introduced. Theterms “host cell” and “recombinant host cell” are used interchangeablyherein. It is understood that such terms refer not only to theparticular subject cell but also to the progeny or potential progeny ofsuch a cell. Because certain modifications may occur in succeedinggenerations due to either mutation or environmental influences, suchprogeny may not, in fact, be identical to the parent cell, but are stillincluded within the scope of the term as used herein.

A host cell can be any prokaryotic or eukaryotic cell. For example,protein of the invention can be produced in bacterial cells such as E.coli, insect cells, yeast, plant or mammalian cells (such as Chinesehamster ovary cells (CHO) or COS or 293 cells). Other suitable hostcells are known to those skilled in the art.

Vector DNA can be introduced into prokaryotic or eukaryotic cells viaconventional transformation or transfection techniques. As used herein,the terms “transformation” and “transfection” are intended to refer to avariety of art-recognized techniques for introducing foreign nucleicacid (e.g., DNA) into a host cell, including calcium phosphate orcalcium chloride co-precipitation, DEAE-dextran-mediated transfection,lipofection, or electroporation. Suitable methods for transforming ortransfecting host cells can be found in Sambrook, et al. (MolecularCloning: A Laboratory Manual. 2nd ed., Cold Spring Harbor Laboratory,Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1989),and other laboratory manuals.

For stable transfection of mammalian cells, it is known that, dependingupon the expression vector and transfection technique used, only a smallfraction of cells may integrate the foreign DNA into their genome. Inorder to identify and select these integrants, a gene that encodes aselectable marker (e.g., resistance to antibiotics) is generallyintroduced into the host cells along with the gene of interest. Variousselectable markers include those that confer resistance to drugs, suchas G418, hygromycin, puromycin, blasticidin and methotrexate. Nucleicacids encoding a selectable marker can be introduced into a host cell onthe same vector as that encoding protein of the invention or can beintroduced on a separate vector. Cells stably transfected with theintroduced nucleic acid can be identified by drug selection (e.g., cellsthat have incorporated the selectable marker gene will survive, whilethe other cells die).

A host cell of the invention, such as a prokaryotic or eukaryotic hostcell in culture, can be used to produce (i.e., express) protein of theinvention. Accordingly, the invention further provides methods forproducing proteins of the invention using the host cells of theinvention. In one embodiment, the method comprises culturing the hostcell of the present invention (into which a recombinant expressionvector encoding protein of the invention has been introduced) in asuitable medium such that the protein of the invention is produced. Inanother embodiment, the method further comprises isolating protein ofthe invention from the medium or the host cell.

For efficient production of the protein, it is preferable to place thenucleotide sequences encoding the protein of the invention under thecontrol of expression control sequences optimized for expression in adesired host. For example, the sequences may include optimizedtranscriptional and/or translational regulatory sequences (such asaltered Kozak sequences).

Protein Modifications

Fusion Proteins

According to the present invention, a fusion protein may be preparedfrom a protein of the invention by fusion with a portion of animmunoglobulin comprising a constant region of an immunoglobulin. Morepreferably, the portion of the immunoglobulin comprises a heavy chainconstant region which is optionally and more preferably a human heavychain constant region. The heavy chain constant region is mostpreferably an IgG heavy chain constant region, and optionally and mostpreferably is an Fc chain, most preferably an IgG Fc fragment thatcomprises CH2 and CH3 domains. Although any IgG subtype may optionallybe used, the IgG1 subtype is preferred. The Fc chain may optionally be aknown or “wild type” Fc chain, or alternatively may be mutated.Non-limiting, illustrative, exemplary types of mutations are describedin US Patent Application No. 20060034852, published on Feb. 16, 2006,hereby incorporated by reference as if fully set forth herein. The term“Fc chain” also optionally comprises any type of Fc fragment.

Several of the specific amino acid residues that are important forantibody constant region-mediated activity in the IgG subclass have beenidentified. Inclusion, substitution or exclusion of these specific aminoacids therefore allows for inclusion or exclusion of specificimmunoglobulin constant region-mediated activity. Furthermore, specificchanges may result in aglycosylation for example and/or other desiredchanges to the Fc chain. At least some changes may optionally be made toblock a function of Fc which is considered to be undesirable, such as anundesirable immune system effect, as described in greater detail below.

Non-limiting, illustrative examples of mutations to Fc which may be madeto modulate the activity of the fusion protein include the followingchanges (given with regard to the Fc sequence nomenclature as given byKabat, from Kabat E A et al: Sequences of Proteins of ImmunologicalInterest. US Department of Health and Human Services, NIH, 1991):220C->S; 233-238 ELLGGP->EAEGAP; 265D->A, preferably in combination with434N->A; 297N->A (for example to block N-glycosylation); 318-322EYKCK->AYACA; 330-331AP->SS; or a combination thereof (see for exampleM. Clark, “Chemical Immunol and Antibody Engineering”, pp 1-31 for adescription of these mutations and their effect). The construct for theFc chain which features the above changes optionally and preferablycomprises a combination of the hinge region with the CH2 and CH3domains.

The above mutations may optionally be implemented to enhance desiredproperties or alternatively to block non-desired properties. Forexample, aglycosylation of antibodies was shown to maintain the desiredbinding functionality while blocking depletion of T-cells or triggeringcytokine release, which may optionally be undesired functions (see M.Clark, “Chemical Immunol and Antibody Engineering”, pp 1-31).Substitution of 331proline for serine may block the ability to activatecomplement, which may optionally be considered an undesired function(see M. Clark, “Chemical Immunol and Antibody Engineering”, pp 1-31).Changing 330alanine to serine in combination with this change may alsoenhance the desired effect of blocking the ability to activatecomplement.

Residues 235 and 237 were shown to be involved in antibody-dependentcell-mediated cytotoxicity (ADCC), such that changing the block ofresidues from 233-238 as described may also block such activity if ADCCis considered to be an undesirable function.

Residue 220 is normally a cysteine for Fc from IgG1, which is the siteat which the heavy chain forms a covalent linkage with the light chain.Optionally, this residue may be changed to a serine, to avoid any typeof covalent linkage (see M. Clark, “Chemical Immunol and AntibodyEngineering”, pp 1-31).

The above changes to residues 265 and 434 may optionally be implementedto reduce or block binding to the Fc receptor, which may optionallyblock undesired functionality of Fc related to its immune systemfunctions (see “Binding site on Human IgG1 for Fc Receptors”, Shields etal, Vol 276, pp 6591-6604, 2001).

The above changes are intended as illustrations only of optional changesand are not meant to be limiting in any way. Furthermore, the aboveexplanation is provided for descriptive purposes only, without wishingto be bound by a single hypothesis.

Addition of Groups

If a protein according to the present invention is a linear molecule, itis possible to place various functional groups at various points on thelinear molecule which are susceptible to or suitable for chemicalmodification. Functional groups can be added to the termini of linearforms of the protein of the invention. In some embodiments, thefunctional groups improve the activity of the protein with regard to oneor more characteristics, including but not limited to, improvement instability, penetration (through cellular membranes and/or tissuebarriers), tissue localization, efficacy, decreased clearance, decreasedtoxicity, improved selectivity, improved resistance to expulsion bycellular pumps, and the like. For convenience sake and without wishingto be limiting, the free N-terminus of one of the sequences contained inthe compositions of the invention will be termed as the N-terminus ofthe composition, and the free C-terminal of the sequence will beconsidered as the C-terminus of the composition. Either the C-terminusor the N-terminus of the sequences, or both, can be linked to acarboxylic acid functional groups or an amine functional group,respectively.

Non-limiting examples of suitable functional groups are described inGreen and Wuts, “Protecting Groups in Organic Synthesis”, John Wiley andSons, Chapters 5 and 7, 1991, the teachings of which are incorporatedherein by reference. Preferred protecting groups are those thatfacilitate transport of the active ingredient attached thereto into acell, for example, by reducing the hydrophilicity and increasing thelipophilicity of the active ingredient, these being an example for “amoiety for transport across cellular membranes”.

These moieties can optionally and preferably be cleaved in vivo, eitherby hydrolysis or enzymatically, inside the cell. (Ditter et al., J.Pharm. Sci. 57:783 (1968); Ditter et al., J. Pharm. Sci. 57:828 (1968);Ditter et al., J. Pharm. Sci. 58:557 (1969); King et al., Biochemistry26:2294 (1987); Lindberg et al., Drug Metabolism and Disposition 17:311(1989); and Tunek et al., Biochem. Pharm. 37:3867 (1988), Anderson etal., Arch. Biochem. Biophys. 239:538 (1985) and Singhal et al., FASEB J.1:220 (1987)). Hydroxyl protecting groups include esters, carbonates andcarbamate protecting groups. Amine protecting groups include alkoxy andaryloxy carbonyl groups, as described above for N-terminal protectinggroups. Carboxylic acid protecting groups include aliphatic, benzylicand aryl esters, as described above for C-terminal protecting groups. Inone embodiment, the carboxylic acid group in the side chain of one ormore glutamic acid or aspartic acid residue in a composition of thepresent invention is protected, preferably with a methyl, ethyl, benzylor substituted benzyl ester, more preferably as a benzyl ester.

Non-limiting, illustrative examples of N-terminal protecting groupsinclude acyl groups (—CO—R1) and alkoxy carbonyl or aryloxy carbonylgroups (−CO—O—R1), wherein R1 is an aliphatic, substituted aliphatic,benzyl, substituted benzyl, aromatic or a substituted aromatic group.Specific examples of acyl groups include but are not limited to acetyl,(ethyl)-CO—, n-propyl-CO—, iso-propyl-CO—, n-butyl-CO—, sec-butyl-CO—,t-butyl-CO—, hexyl, lauroyl, palmitoyl, myristoyl, stearyl, oleoylphenyl-CO—, substituted phenyl-CO—, benzyl-CO— and (substitutedbenzyl)-CO—. Examples of alkoxy carbonyl and aryloxy carbonyl groupsinclude CH3-O—CO—, (ethyl)-O—CO—, n-propyl-O—CO—, iso-propyl-O—CO—,n-butyl-O—CO—, sec-butyl-O—CO—, t-butyl-O—CO—, phenyl-O—CO—, substitutedphenyl-O—CO— and benzyl-O—CO—, (substituted benzyl)-O—CO—, Adamantan,naphtalen, myristoleyl, toluen, biphenyl, cinnamoyl, nitrobenzoy,toluoyl, furoyl, benzoyl, cyclohexane, norbornane, or Z-caproic. Inorder to facilitate the N-acylation, one to four glycine residues can bepresent in the N-terminus of the molecule.

The carboxyl group at the C-terminus of the compound can be protected,for example, by a group including but not limited to an amide (i.e., thehydroxyl group at the C-terminus is replaced with —NH₂, —NHR₂ and—NR₂R₃) or ester (i.e. the hydroxyl group at the C-terminus is replacedwith —OR₂). R₂ and R₃ are optionally independently an aliphatic,substituted aliphatic, benzyl, substituted benzyl, aryl or a substitutedaryl group. In addition, taken together with the nitrogen atom, R₂ andR₃ can optionally form a C4 to C8 heterocyclic ring with from about 0-2additional heteroatoms such as nitrogen, oxygen or sulfur. Non-limitingsuitable examples of suitable heterocyclic rings include piperidinyl,pyrrolidinyl, morpholino, thiomorpholino or piperazinyl. Examples ofC-terminal protecting groups include but are not limited to —NH₂,—NHCH₃, —N(CH₃)₂, —NH(ethyl), —N(ethyl)₂, —N(methyl)(ethyl),—NH(benzyl), —N(C1-C4 alkyl)(benzyl), —NH(phenyl), —N(C1-C4 alkyl)(phenyl), —OCH₃, —O-(ethyl), —O-(n-propyl), —O-(n-butyl),—O-(iso-propyl), —O-(sec-butyl), —O-(t-butyl), —O-benzyl and —O-phenyl.

Substitution by Peptidomimetic Moieties

A “peptidomimetic organic moiety” can optionally be substituted foramino acid residues in the composition of this invention both asconservative and as non-conservative substitutions. These moieties arealso termed “non-natural amino acids” and may optionally replace aminoacid residues, amino acids or act as spacer groups within the peptidesin lieu of deleted amino acids. The peptidomimetic organic moietiesoptionally and preferably have steric, electronic or configurationalproperties similar to the replaced amino acid and such peptidomimeticsare used to replace amino acids in the essential positions, and areconsidered conservative substitutions. However such similarities are notnecessarily required. According to preferred embodiments of the presentinvention, one or more peptidomimetics are selected such that thecomposition at least substantially retains its physiological activity ascompared to the native protein according to the present invention.

Peptidomimetics may optionally be used to inhibit degradation of thepeptides by enzymatic or other degradative processes. Thepeptidomimetics can optionally and preferably be produced by organicsynthetic techniques. Non-limiting examples of suitable peptidomimeticsinclude D amino acids of the corresponding L amino acids, tetrazol(Zabrocki et al., J. Am. Chem. Soc. 110:5875-5880 (1988)); isosteres ofamide bonds (Jones et al., Tetrahedron Lett. 29: 3853-3856 (1988));LL-3-amino-2-propenidone-6-carboxylic acid (LL-Acp) (Kemp et al., J.Org. Chem. 50:5834-5838 (1985)). Similar analogs are shown in Kemp etal., Tetrahedron Lett. 29:5081-5082 (1988) as well as Kemp et al.,Tetrahedron Lett. 29:5057-5060 (1988), Kemp et al., Tetrahedron Lett.29:4935-4938 (1988) and Kemp et al., J. Org. Chem. 54:109-115 (1987).Other suitable but exemplary peptidomimetics are shown in Nagai andSato, Tetrahedron Lett. 26:647-650 (1985); Di Maio et al., J. Chem. Soc.Perkin Trans., 1687 (1985); Kahn et al., Tetrahedron Lett. 30:2317(1989); Olson et al., J. Am. Chem. Soc. 112:323-333 (1990); Garvey etal., J. Org. Chem. 56:436 (1990). Further suitable exemplarypeptidomimetics includehydroxy-1,2,3,4-tetrahydroisoquinoline-3-carboxylate (Miyake et al., J.Takeda Res. Labs 43:53-76 (1989));1,2,3,4-tetrahydro-isoquinoline-3-carboxylate (Kazmierski et al., J. Am.Chem. Soc. 133:2275-2283 (1991)); histidine isoquinolone carboxylic acid(HIC) (Zechel et al., Int. J. Pep. Protein Res. 43 (1991));(2S,3S)-methyl-phenylalanine, (2S, 3R)-methyl-phenylalanine,(2R,3S)-methyl-phenylalanine and (2R, 3R)-methyl-phenylalanine(Kazmierski and Hruby, Tetrahedron Lett. (1991)).

Exemplary, illustrative but non-limiting non-natural amino acids includebeta-amino acids (beta3 and beta2), homo-amino acids, cyclic aminoacids, aromatic amino acids, Pro and Pyr derivatives, 3-substitutedAlanine derivatives, Glycine derivatives, ring-substituted Phe and TyrDerivatives, linear core amino acids or diamino acids. They areavailable from a variety of suppliers, such as Sigma-Aldrich (USA) forexample.

Chemical Modifications

In the present invention any part of a protein of the invention mayoptionally be chemically modified, i.e. changed by addition offunctional groups. For example the side amino acid residues appearing inthe native sequence may optionally be modified, although as describedbelow alternatively other parts of the protein may optionally bemodified, in addition to or in place of the side amino acid residues.The modification may optionally be performed during synthesis of themolecule if a chemical synthetic process is followed, for example byadding a chemically modified amino acid. However, chemical modificationof an amino acid when it is already present in the molecule (“in situ”modification) is also possible.

The amino acid of any of the sequence regions of the molecule canoptionally be modified according to any one of the following exemplarytypes of modification (in the peptide conceptually viewed as “chemicallymodified”). Non-limiting exemplary types of modification includecarboxymethylation, acylation, phosphorylation, glycosylation or fattyacylation. Ether bonds can optionally be used to join the serine orthreonine hydroxyl to the hydroxyl of a sugar. Amide bonds canoptionally be used to join the glutamate or aspartate carboxyl groups toan amino group on a sugar (Garg and Jeanloz, Advances in CarbohydrateChemistry and Biochemistry, Vol. 43, Academic Press (1985); Kunz, Ang.Chem. Int. Ed. English 26:294-308 (1987)). Acetal and ketal bonds canalso optionally be formed between amino acids and carbohydrates. Fattyacid acyl derivatives can optionally be made, for example, by acylationof a free amino group (e.g., lysine) (Toth et al., Peptides: Chemistry,Structure and Biology, Rivier and Marshal, eds., ESCOM Publ., Leiden,1078-1079 (1990)).

As used herein the term “chemical modification”, when referring to aprotein or peptide according to the present invention, refers to aprotein or peptide where at least one of its amino acid residues ismodified either by natural processes, such as processing or otherpost-translational modifications, or by chemical modification techniqueswhich are well known in the art. Examples of the numerous knownmodifications typically include, but are not limited to: acetylation,acylation, amidation, ADP-ribosylation, glycosylation, GPI anchorformation, covalent attachment of a lipid or lipid derivative,methylation, myristylation, pegylation, prenylation, phosphorylation,ubiquitination, or any similar process.

Other types of modifications optionally include the addition of acycloalkane moiety to a biological molecule, such as a protein, asdescribed in PCT Application No. WO 2006/050262, hereby incorporated byreference as if fully set forth herein. These moieties are designed foruse with biomolecules and may optionally be used to impart variousproperties to proteins.

Furthermore, optionally any point on a protein may be modified. Forexample, pegylation of a glycosylation moiety on a protein mayoptionally be performed, as described in PCT Application No. WO2006/050247, hereby incorporated by reference as if fully set forthherein. One or more polyethylene glycol (PEG) groups may optionally beadded to O-linked and/or N-linked glycosylation. The PEG group mayoptionally be branched or linear. Optionally any type of water-solublepolymer may be attached to a glycosylation site on a protein through aglycosyl linker.

Altered Glycosylation

Proteins of the invention may be modified to have an alteredglycosylation pattern (i.e., altered from the original or nativeglycosylation pattern). As used herein, “altered” means having one ormore carbohydrate moieties deleted, and/or having at least oneglycosylation site added to the original protein.

Glycosylation of proteins is typically either N-linked or O-linked.N-linked refers to the attachment of the carbohydrate moiety to the sidechain of an asparagine residue. The tripeptide sequences,asparagine-X-serine and asparagine-X-threonine, where X is any aminoacid except proline, are the recognition sequences for enzymaticattachment of the carbohydrate moiety to the asparagine side chain.Thus, the presence of either of these tripeptide sequences in apolypeptide creates a potential glycosylation site. O-linkedglycosylation refers to the attachment of one of the sugarsN-acetylgalactosamine, galactose, or xylose to a hydroxyamino acid, mostcommonly serine or threonine, although 5-hydroxyproline or5-hydroxylysine may also be used.

Addition of glycosylation sites to proteins of the invention isconveniently accomplished by altering the amino acid sequence of theprotein such that it contains one or more of the above-describedtripeptide sequences (for N-linked glycosylation sites). The alterationmay also be made by the addition of, or substitution by, one or moreserine or threonine residues in the sequence of the original protein(for O-linked glycosylation sites). The protein's amino acid sequencemay also be altered by introducing changes at the DNA level.

Another means of increasing the number of carbohydrate moieties onproteins is by chemical or enzymatic coupling of glycosides to the aminoacid residues of the protein. Depending on the coupling mode used, thesugars may be attached to (a) arginine and histidine, (b) free carboxylgroups, (c) free sulfhydryl groups such as those of cysteine, (d) freehydroxyl groups such as those of serine, threonine, or hydroxyproline,(e) aromatic residues such as those of phenylalanine, tyrosine, ortryptophan, or (f) the amide group of glutamine. These methods aredescribed in WO 87/05330, and in Aplin and Wriston, CRC Crit. Rev.Biochem., 22: 259-306 (1981).

Removal of any carbohydrate moieties present on proteins of theinvention may be accomplished chemically or enzymatically. Chemicaldeglycosylation requires exposure of the protein totrifluoromethanesulfonic acid, or an equivalent compound. This treatmentresults in the cleavage of most or all sugars except the linking sugar(N-acetylglucosamine or N-acetylgalactosamine), leaving the amino acidsequence intact.

Chemical deglycosylation is described by Hakimuddin et al., Arch.Biochem. Biophys., 259: 52 (1987); and Edge et al., Anal. Biochem., 118:131 (1981). Enzymatic cleavage of carbohydrate moieties on proteins canbe achieved by the use of a variety of endo- and exo-glycosidases asdescribed by Thotakura et al., Meth. Enzymol., 138: 350 (1987).

Methods of Treatment

As mentioned hereinabove the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32,FXYD3 proteins or VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3proteins and polypeptides of the present invention or nucleic acidsequence or fragments thereof especially the ectodomain or secretedforms of VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 proteins, aswell as drugs which specifically bind to the VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, FXYD3 proteins and/or splice variants, and/or drugswhich agonize or antagonize the binding of other moieties to the VSIG1,ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 proteins and/or splicevariants, and/or drugs which modulate (agonize or antagonize) at leastone VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 relatedbiological activity (such drugs include by way of example antibodies,small molecules, peptides, ribozymes, antisense molecules, siRNA's andthe like), can be used to treat cancer, including but not limited tonon-solid and solid tumors, sarcomas, hematological malignanciesincluding but not limited to acute lymphocytic leukemia, chroniclymphocytic leukemia, acute myelogenous leukemia, chronic myelogenousleukemia, multiple myeloma, Hodgkin's lymphoma, Non-Hodgkin's lymphoma,cancer of the breast, prostate, lung, ovary, colon, spleen, kidney,bladder, head and neck, uterus, testicles, stomach, cervix, liver, bone,skin, pancreas, brain and wherein the cancer may be non-metastatic,invasive or metastatic.

The VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 proteins or VSIG1,ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 proteins and polypeptides ofthe present invention or nucleic acid sequence or fragments thereofespecially the ectodomain or secreted forms of VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, FXYD3 proteins, as well as drugs which specificallybind to the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 proteinsand/or splice variants, and/or drugs which agonize or antagonize thebinding of other moieties to the VSIG1, ILDR1, LOC253012, AI216611,C1ORF32, FXYD3 proteins and/or splice variants, and/or drugs whichmodulate (agonize or antagonize) at least one VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 or FXYD3 related biological activity (such drugsinclude by way of example antibodies, small molecules, peptides,ribozymes, antisense molecules, siRNA's and the like), can be furtherused to treat non-malignant disorders such as immune disorders includingbut not limited to autoimmune diseases, transplant rejection and graftversus host disease, and/or for blocking or promoting immunecostimulation mediated by the VSIG1, ILDR1, LOC253012, AI216611,C1ORF32, FXYD3 polypeptide.

Thus, according to an additional aspect of the present invention thereis provided a method of treating cancer, including but not limited tonon-solid and solid tumors, sarcomas, hematological malignanciesincluding but not limited to acute lymphocytic leukemia, chroniclymphocytic leukemia, acute myelogenous leukemia, chronic myelogenousleukemia, multiple myeloma, Hodgkin's lymphoma, Non-Hodgkin's lymphoma,cancer of the breast, prostate, lung, ovary, colon, spleen, kidney,bladder, head and neck, uterus, testicles, stomach, cervix, liver, bone,skin, pancreas, brain and wherein the cancer may be non-metastatic,invasive or metastatic as well as non-malignant disorders such as immunedisorders including but not limited to autoimmune diseases, transplantrejection and graft versus host disease, and/or for blocking orpromoting immune costimulation mediated by the VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, or FXYD3 polypeptide in a subject.

The subject according to the present invention is a mammal, preferably ahuman which is diagnosed with one of the disease, disorder or conditionsdescribed hereinabove, or alternatively is predisposed to at least onetype of cancer, including but not limited to non-solid and solid tumors,sarcomas, hematological malignancies including but not limited to acutelymphocytic leukemia, chronic lymphocytic leukemia, acute myelogenousleukemia, chronic myelogenous leukemia, multiple myeloma, Hodgkin'slymphoma, Non-Hodgkin's lymphoma, cancer of the breast, prostate, lung,ovary, colon, spleen, kidney, bladder, head and neck, uterus, testicles,stomach, cervix, liver, bone, skin, pancreas, brain, as well asnon-malignant disorders such as immune disorders including but notlimited to autoimmune diseases, transplant rejection and graft versushost disease.

As used herein the term “treating” refers to preventing, curing,reversing, attenuating, alleviating, minimizing, suppressing or haltingthe deleterious effects of the above-described diseases, disorders orconditions.

Treating, according to the present invention, can be effected byspecifically upregulating the expression of at least one of thepolypeptides of the present invention in the subject.

Optionally, upregulation may be effected by administering to the subjectat least one of the polypeptides of the present invention (e.g.,recombinant or synthetic) or an active portion thereof, as describedherein. However, since the bioavailability of large polypeptides maypotentially be relatively small due to high degradation rate and lowpenetration rate, administration of polypeptides is preferably confinedto small peptide fragments (e.g., about 100 amino acids). Thepolypeptide or peptide may optionally be administered in as part of apharmaceutical composition, described in more detail below.

It will be appreciated that treatment of the above-described diseasesaccording to the present invention may be combined with other treatmentmethods known in the art (i.e., combination therapy). Thus, treatment ofmalignancies using the agents of the present invention may be combinedwith, for example, radiation therapy, antibody therapy and/orchemotherapy.

Alternatively or additionally, an upregulating method may optionally beeffected by specifically upregulating the amount (optionally expression)in the subject of at least one of the polypeptides of the presentinvention or active portions thereof.

As is mentioned hereinabove and in the Examples section which follows,the biomolecular sequences of this aspect of the present invention maybe used as valuable therapeutic tools in the treatment of diseases,disorders or conditions in which altered activity or expression of thewild-type gene product (known protein) is known to contribute todisease, disorder or condition onset or progression. For example, incase a disease is caused by overexpression of a membrane bound-receptor,a soluble variant thereof may be used as an antagonist which competeswith the receptor for binding the ligand, to thereby terminate signalingfrom the receptor.

Anti-VSIG1, Anti-ILDR1, Anti-LOC253012, Anti-AI216611, Anti-C1ORF32,Anti-FXYD3 Antibodies

The antibodies of the invention including those having the particulargermline sequences, homologous antibodies, antibodies with conservativemodifications, engineered and modified antibodies are characterized byparticular functional features or properties of the antibodies. Forexample, the antibodies bind specifically to human VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, or FXYD3. Preferably, an antibody of theinvention binds to corresponding VSIG1, ILDR1, LOC253012, AI216611,C1ORF32, or FXYD3 with high affinity, for example with a KD of 10−8 M orless or 10−9 M or less or even 10−10 M or less. The anti-VSIG1,anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32, or anti-FXYD3antibodies of the invention preferably exhibit one or more of thefollowing characteristics:

(i) binds to corresponding human VSIG1, ILDR1, LOC253012, AI216611,C1ORF32, or FXYD3 with a KD of 5×10−8 M or less;

(ii) modulates (enhances or inhibits) B7 immune costimulation andrelated activities and functions such a T cell responses involved inantitumor immunity and autoimmunity. and/or

(iii) binds to VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3antigen expressed by cancer cells including for example lung cancer,ovarian cancer, colon cancer, but does not substantially bind to normalcells In addition, preferably these antibodies and conjugates thereofwill be effective in eliciting selective killing of such cancer cellsand for modulating immune responses involved in autoimmunity and cancer.

More preferably, the antibody binds to corresponding human VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, or FXYD3 antigen with a KD of 3×10−8 M orless, or with a KD of 1×10−9 M or less, or with a KD of 0.1×10−9 M orless, or with a KD Of 0.05×10−9 M or less or with a KD of between 1×10−9and 1×10−11 M.

Standard assays to evaluate the binding ability of the antibodies towardVSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3 are known in theart, including for example, ELISAs, Western blots and RIAs. Suitableassays are described in detail in the Examples. The binding kinetics(e.g., binding affinity) of the antibodies also can be assessed bystandard assays known in the art, such as by Biacore analysis.

Upon production of anti-VSIG1, anti-ILDR1, anti-LOC253012,anti-AI216611, anti-C1ORF32, or anti-FXYD3 antibody sequences fromantibodies can bind to VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, orFXYD3 the VH and VL sequences can be “mixed and matched” to create otheranti-VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3 bindingmolecules of the invention. VSIG1, ILDR1, LOC253012, AI216611, C1ORF32,or FXYD3 binding of such “mixed and matched” antibodies can be testedusing the binding assays described above. e.g., ELISAs). Preferably,when VH and VL chains are mixed and matched, a VH sequence from aparticular VH/VL pairing is replaced with a structurally similar VHsequence. Likewise, preferably a VL sequence from a particular VH/VLpairing is replaced with a structurally similar VL sequence. Forexample, the VH and VL sequences of homologous antibodies areparticularly amenable for mixing and matching.

Antibodies Having Particular Germline Sequences

In certain embodiments, an antibody of the invention comprises a heavychain variable region from a particular germline heavy chainimmunoglobulin gene and/or a light chain variable region from aparticular germline light chain immunoglobulin gene.

As used herein, a human antibody comprises heavy or light chain variableregions that is “the product of” or “derived from” a particular germlinesequence if the variable regions of the antibody are obtained from asystem that uses human germline immunoglobulin genes. Such systemsinclude immunizing a transgenic mouse carrying human immunoglobulingenes with the antigen of interest or screening a human immunoglobulingene library displayed on phage with the antigen of interest. A humanantibody that is “the product” of or “derived from” a human germlineimmunoglobulin sequence can be identified as such by comparing the aminoacid sequence of the human antibody to the amino acid sequences of humangermline immunoglobulins and selecting the human germline immunoglobulinsequence that is closest in sequence (i.e., greatest % identity) to thesequence of the human antibody.

A human antibody that is “the product of” or “derived from” a particularhuman germline immunoglobulin sequence may contain amino aciddifferences as compared to the germline sequence, due to, for example,naturally-occurring somatic mutations or intentional introduction ofsite-directed mutation. However, a selected human antibody typically isat least 90% identical in amino acids sequence to an amino acid sequenceencoded by a human germline immunoglobulin gene and contains amino acidresidues that identify the human antibody as being human when comparedto the germline immunoglobulin amino acid sequences of other species(e.g., murine germline sequences). In certain cases, a human antibodymay be at least 95, 96, 97, 98 or 99%, or even at least 96%, 97%, 98%,or 99% identical in amino acid sequence to the amino acid sequenceencoded by the germline immunoglobulin gene. Typically, a human antibodyderived from a particular human germline sequence will display no morethan 10 amino acid differences from the amino acid sequence encoded bythe human germline immunoglobulin gene. In certain cases, the humanantibody may display no more than 5, or even no more than 4, 3, 2, or 1amino acid difference from the amino acid sequence encoded by thegermline immunoglobulin gene.

Homologous Antibodies

In yet another embodiment, an antibody of the invention comprises heavyand light chain variable regions comprising amino acid sequences thatare homologous to isolated anti-VSIG1, anti-ILDR1, anti-LOC253012,anti-AI216611, anti-C1ORF32, or anti-FXYD3 amino acid sequences ofpreferred anti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611,anti-C1ORF32, or anti-FXYD3 antibodies, respectively, wherein theantibodies retain the desired functional properties of the parentanti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32, oranti-FXYD3 antibodies.

As used herein, the percent homology between two amino acid sequences isequivalent to the percent identity between the two sequences. Thepercent identity between the two sequences is a function of the numberof identical positions shared by the sequences (i.e., % homology=# ofidentical positions/total # of positions×100), taking into account thenumber of gaps, and the length of each gap, which need to be introducedfor optimal alignment of the two sequences. The comparison of sequencesand determination of percent identity between two sequences can beaccomplished using a mathematical algorithm, as described in thenon-limiting examples below.

The percent identity between two amino acid sequences can be determinedusing the algorithm of E. Meyers and W. Miller (Comput. Appl. Biosci.,4:11-17 (1988)) which has been incorporated into the ALIGN program(version 2.0), using a PAM120 weight residue table, a gap length penaltyof 12 and a gap penalty of 4. In addition, the percent identity betweentwo amino acid sequences can be determined using the Needleman andWunsch (J. Mol. Biol. 48:444-453 (1970)) algorithm which has beenincorporated into the GAP program in the GCG software package (availablecommercially), using either a Blossum 62 matrix or a PAM250 matrix, anda gap weight of 16, 14, 12, 10, 8, 6, or 4 and a length weight of 1, 2,3, 4, 5, or 6.

Additionally or alternatively, the protein sequences of the presentinvention can further be used as a “query sequence” to perform a searchagainst public databases to, for example, identify related sequences.Such searches can be performed using the XBLAST program (version 2.0) ofAltschul, et al. (1990) J Mol. Biol. 215:403-10. BLAST protein searchescan be performed with the XBLAST program, score=50, wordlength=3 toobtain amino acid sequences homologous to the antibody molecules of theinvention. To obtain gapped alignments for comparison purposes, GappedBLAST can be utilized as described in Altschul et al., (1997) NucleicAcids Res. 25(17):3389-3402. When utilizing BLAST and Gapped BLASTprograms, the default parameters of the respective programs (e.g.,XBLAST and NBLAST) can be used.

Antibodies with Conservative Modifications

In certain embodiments, an antibody of the invention comprises a heavychain variable region comprising CDR1, CDR2 and CDR3 sequences and alight chain variable region comprising CDR1, CDR2 and CDR3 sequences,wherein one or more of these CDR sequences comprise specified amino acidsequences based on preferred anti-VSIG1, anti-ILDR1, anti-LOC253012,anti-AI216611, anti-C1ORF32, or anti-FXYD3 antibodies isolated andproduced using methods herein, or conservative modifications thereof,and wherein the antibodies retain the desired functional properties ofthe anti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32,or anti-FXYD3 antibodies of the invention, respectively.

In various embodiments, the anti-VSIG1, anti-ILDR1, anti-LOC253012,anti-AI216611, anti-C1ORF32, or anti-FXYD3 antibody can be, for example,human antibodies, humanized antibodies or chimeric antibodies.

As used herein, the term “conservative sequence modifications” isintended to refer to amino acid modifications that do not significantlyaffect or alter the binding characteristics of the antibody containingthe amino acid sequence. Such conservative modifications include aminoacid substitutions, additions and deletions. Modifications can beintroduced into an antibody of the invention by standard techniquesknown in the art, such as site-directed mutagenesis and PCR-mediatedmutagenesis. Conservative amino acid substitutions are ones in which theamino acid residue is replaced with an amino acid residue having asimilar side chain. Families of amino acid residues having similar sidechains have been defined in the art. These families include amino acidswith basic side chains (e.g., lysine, arginine, histidine), acidic sidechains (e.g., aspartic acid, glutamic acid), uncharged polar side chains(e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine,cysteine, tryptophan), nonpolar side chains (e.g., alanine, valine,leucine, isoleucine, proline, phenylalanine, methionine), beta-branchedside chains (e.g., threonine, valine, isoleucine) and aromatic sidechains (e.g., tyrosine, phenylalanine, tryptophan, histidine). Thus, oneor more amino acid residues within the CDR regions of an antibody of theinvention can be replaced with other amino acid residues from the sameside chain family and the altered antibody can be tested for retainedfunction (i.e., the functions set forth in (c) through (j) above) usingthe functional assays described herein.

Antibodies that Bind to the Same Epitope as Anti-VSIG1, Anti-ILDR1,Anti-LOC253012, Anti-AI216611, Anti-C1ORF32, or Anti-FXYD3 Antibodies ofthe Invention

In another embodiment, the invention provides antibodies that bind topreferred epitopes on human VSIG1, ILDR1, LOC253012, AI216611, C1ORF32,or FXYD3 which possess desired functional properties such as modulationof B7 co-stimulation and related functions. Other antibodies withdesired epitope specificity may be selected and will have the ability tocross-compete for binding to VSIG1, ILDR1, LOC253012, AI216611, C1ORF32,or FXYD3 antigen with the desired antibodies.

Engineered and Modified Antibodies

An antibody of the invention further can be prepared using an antibodyhaving one or more of the VH and/or VL sequences derived from ananti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32, oranti-FXYD3 antibody starting material to engineer a modified antibody,which modified antibody may have altered properties from the startingantibody. An antibody can be engineered by modifying one or moreresidues within one or both variable regions (i.e., VH and/or VL), forexample within one or more CDR regions and/or within one or moreframework regions. Additionally or alternatively, an antibody can beengineered by modifying residues within the constant regions, forexample to alter the effector functions of the antibody.

One type of variable region engineering that can be performed is CDRgrafting. Antibodies interact with target antigens predominantly throughamino acid residues that are located in the six heavy and light chaincomplementarity determining regions (CDRs). For this reason, the aminoacid sequences within CDRs are more diverse between individualantibodies than sequences outside of CDRs. Because CDR sequences areresponsible for most antibody-antigen interactions, it is possible toexpress recombinant antibodies that mimic the properties of specificnaturally occurring antibodies by constructing expression vectors thatinclude CDR sequences from the specific naturally occurring antibodygrafted onto framework sequences from a different antibody withdifferent properties (see, e.g., Riechmann, L. et al. (1998) Nature332:323-327; Jones, P. et al. (1986) Nature 321:522-525; Queen, C. etal. (1989) Proc. Natl. Acad. See. U.S.A. 86:10029-10033; U.S. Pat. No.5,225,539 to Winter, and U.S. Pat. Nos. 5,530,101; 5,585,089; 5,693,762and 6,180,370 to Queen et al.)

Suitable framework sequences can be obtained from public DNA databasesor published references that include germline antibody gene sequences.For example, germline DNA sequences for human heavy and light chainvariable region genes can be found in the “VBase” human germlinesequence database (available on the Internet), as well as in Kabat, E.A., et al. (1991) Sequences of Proteins of Immunological Interest, FifthEdition, U.S. Department of Health and Human Services, NIH PublicationNo. 91-3242; Tomlinson, I. M., et al. (1992) “The Repertoire of HumanGermline VH Sequences Reveals about Fifty Groups of VH Segments withDifferent Hypervariable Loops” J. Mol. Biol. 227:776-798; and Cox, J. P.L. et al. (1994) “A Directory of Human Germ-line VH Segments Reveals aStrong Bias in their Usage” Eur. J. Immunol. 24:827-836; the contents ofeach of which are expressly incorporated herein by reference.

Another type of variable region modification is to mutate amino acidresidues within the VH and/or VL CDR 1, CDR2 and/or CDR3 regions tothereby improve one or more binding properties (e.g., affinity) of theantibody of interest. Site-directed mutagenesis or PCR-mediatedmutagenesis can be performed to introduce the mutations and the effecton antibody binding, or other functional property of interest, can beevaluated in appropriate in vitro or in vivo assays. Preferablyconservative modifications (as discussed above) are introduced. Themutations may be amino acid substitutions, additions or deletions, butare preferably substitutions. Moreover, typically no more than one, two,three, four or five residues within a CDR region are altered.

Engineered antibodies of the invention include those in whichmodifications have been made to framework residues within VH and/or VL,e.g. to improve the properties of the antibody. Typically such frameworkmodifications are made to decrease the immunogenicity of the antibody.For example, one approach is to “backmutate” one or more frameworkresidues to the corresponding germline sequence. More specifically, anantibody that has undergone somatic mutation may contain frameworkresidues that differ from the germline sequence from which the antibodyis derived. Such residues can be identified by comparing the antibodyframework sequences to the germline sequences from which the antibody isderived.

In addition or alternative to modifications made within the framework orCDR regions, antibodies of the invention may be engineered to includemodifications within the Fc region, typically to alter one or morefunctional properties of the antibody, such as serum half-life,complement fixation, Fc receptor binding, and/or antigen-dependentcellular cytotoxicity. Furthermore, an antibody of the invention may bechemically modified (e.g., one or more chemical moieties can be attachedto the antibody) or be modified to alter its glycosylation, again toalter one or more functional properties of the antibody. Suchembodiments are described further below. The numbering of residues inthe Fc region is that of the EU index of Kabat.

In one embodiment, the hinge region of CH1 is modified such that thenumber of cysteine residues in the hinge region is altered, e.g.,increased or decreased. This approach is described further in U.S. Pat.No. 5,677,425 by Bodmer et al. The number of cysteine residues in thehinge region of CH1 is altered to, for example, facilitate assembly ofthe light and heavy chains or to increase or decrease the stability ofthe antibody.

In another embodiment, the Fc hinge region of an antibody is mutated todecrease the biological half life of the antibody. More specifically,one or more amino acid mutations are introduced into the CH2-CH3 domaininterface region of the Fc-hinge fragment such that the antibody hasimpaired Staphylococcyl protein A (SpA) binding relative to nativeFc-hinge domain SpA binding. This approach is described in furtherdetail in U.S. Pat. No. 6,165,745 by Ward et al.

In another embodiment, the antibody is modified to increase itsbiological half life. Various approaches are possible. For example, oneor more of the following mutations can be introduced: T252L, T254S,T256F, as described in U.S. Pat. No. 6,277,375 to Ward. Alternatively,to increase the biological half life, the antibody can be altered withinthe CH1 or CL region to contain a salvage receptor binding epitope takenfrom two loops of a CH2 domain of an Fc region of an IgG, as describedin U.S. Pat. Nos. 5,869,046 and 6,121,022 by Presta et al.

In yet other embodiments, the Fc region is altered by replacing at leastone amino acid residue with a different amino acid residue to alter theeffector functions of the antibody. For example, one or more amino acidsselected from amino acid residues 234, 235, 236, 237, 297, 318, 320 and322 can be replaced with a different amino acid residue such that theantibody has an altered affinity for an effector ligand but retains theantigen-binding ability of the parent antibody. The effector ligand towhich affinity is altered can be, for example, an Fc receptor or the C1component of complement. This approach is described in further detail inU.S. Pat. Nos. 5,624,821 and 5,648,260, both by Winter et al.

In another example, one or more amino acids selected from amino acidresidues 329, 331 and 322 can be replaced with a different amino acidresidue such that the antibody has altered C1q binding and/or reduced orabolished complement dependent cytotoxicity (CDC). This approach isdescribed in further detail in U.S. Pat. No. 6,194,551 by Idusogie etal.

In another example, one or more amino acid residues within amino acidpositions 231 and 239 are altered to thereby alter the ability of theantibody to fix complement. This approach is described further in PCTPublication WO 94/29351 by Bodmer et al.

In yet another example, the Fc region is modified to increase theability of the antibody to mediate antibody dependent cellularcytotoxicity (ADCC) and/or to increase the affinity of the antibody foran Fcy receptor by modifying one or more amino acids at the followingpositions: 238, 239, 248, 249, 252, 254, 255, 256, 258, 265, 267, 268,269, 270, 272, 276, 278, 280, 283, 285, 286, 289, 290, 292, 293, 294,295, 296, 298, 301, 303, 305, 307, 309, 312, 315, 320, 322, 324, 326,327, 329, 330, 331, 333, 334, 335, 337, 338, 340, 360, 373, 376, 378,382, 388, 389, 398, 414, 416, 419, 430, 434, 435, 437, 438 or 439. Thisapproach is described further in PCT Publication WO 00/42072 by Presta.Moreover, the binding sites on human IgG1 for Fc grammar, Fc gamma RII,Fc gammaRIII and FcRn have been mapped and variants with improvedbinding have been described (see Shields, R. L. et al. (2001) J. Biol.Chem. 276:6591-6604). Specific mutations at positions 256, 290, 298,333, 334 and 339 are shown to improve binding to FcyRIII. Additionally,the following combination mutants are shown to improve Fcgamma.RIIIbinding: T256A/S298A, S298A/E333A, S298A/K224A and S298A/E333A/K334A.

In still another embodiment, the glycosylation of an antibody ismodified. For example, an aglycoslated antibody can be made (i.e., theantibody lacks glycosylation). Glycosylation can be altered to, forexample, increase the affinity of the antibody for antigen. Suchcarbohydrate modifications can be accomplished by, for example, alteringone or more sites of glycosylation within the antibody sequence. Forexample, one or more amino acid substitutions can be made that result inelimination of one or more variable region framework glycosylation sitesto thereby eliminate glycosylation at that site. Such aglycosylation mayincrease the affinity of the antibody for antigen. Such an approach isdescribed in further detail in U.S. Pat. Nos. 5,714,350 and 6,350,861 byCo et al.

Additionally or alternatively, an antibody can be made that has analtered type of glycosylation, such as a hypofucosylated antibody havingreduced amounts of fucosyl residues or an antibody having increasedbisecting GlcNac structures. Such altered glycosylation patterns havebeen demonstrated to increase the ADCC ability of antibodies. Suchcarbohydrate modifications can be accomplished by, for example,expressing the antibody in a host cell with altered glycosylationmachinery. Cells with altered glycosylation machinery have beendescribed in the art and can be used as host cells in which to expressrecombinant antibodies of the invention to thereby produce an antibodywith altered glycosylation. For example, the cell lines Ms704, Ms705,and Ms709 lack the fucosyltransferase gene, FUT8(alpha(1,6)fucosyltransferase), such that antibodies expressed in theMs704, Ms705, and Ms709 cell lines lack fucose on their carbohydrates.The Ms704, Ms705, and Ms709 FUT8.−/− cell lines are created by thetargeted disruption of the FUT8 gene in CHO/DG44 cells using tworeplacement vectors (see U.S. Patent Publication No. 20040110704 byYamane et al. and Yamane-Ohnuki et al. (2004) Biotechnol Bioeng87:614-22). As another example, EP 1,176,195 by Hanai et al. describes acell line with a functionally disrupted FUT8 gene, which encodes afucosyl transferase, such that antibodies expressed in such a cell lineexhibit hypofucosylation by reducing or eliminating the alpha 1,6bond-related enzyme. Hanai et al. also describe cell lines which have alow enzyme activity for adding fucose to the N-acetylglucosamine thatbinds to the Fc region of the antibody or does not have the enzymeactivity, for example the rat myeloma cell line YB2/0 (ATCC CRL 1662).PCT Publication WO 03/035835 by Presta describes a variant CHO cellline, Lec13 cells, with reduced ability to attach fucose toAsn(297)-linked carbohydrates, also resulting in hypofucosylation ofantibodies expressed in that host cell (see also Shields, R. L. et al.(2002) J. Biol. Chem. 277:26733-26740). PCT Publication WO 99/54342 byUmana et al. describes cell lines engineered to expressglycoprotein-modifying glycosyl transferases (e.g.,beta(1,4)-N-acetylglucosaminyltransferase III (GnTIII)) such thatantibodies expressed in the engineered cell lines exhibit increasedbisecting GlcNac structures which results in increased ADCC activity ofthe antibodies (see also Umana et al. (1999) Nat. Biotech. 17:176-180).Alternatively, the fucose residues of the antibody may be cleaved offusing a fucosidase enzyme. For example, the fucosidasealpha-L-fucosidase removes fucosyl residues from antibodies (Tarentino,A. L. et al. (1975) Biochem. 14:5516-23).

Another modification of the antibodies herein that is contemplated bythe invention is pegylation. An antibody can be pegylated to, forexample, increase the biological (e.g., serum) half life of theantibody. To pegylate an antibody, the antibody, or fragment thereof,typically is reacted with polyethylene glycol (PEG), such as a reactiveester or aldehyde derivative of PEG, under conditions in which one ormore PEG groups become attached to the antibody or antibody fragment.Preferably, the pegylation is carried out via an acylation reaction oran alkylation reaction with a reactive PEG molecule (or an analogousreactive water-soluble polymer). As used herein, the term “polyethyleneglycol” is intended to encompass any of the forms of PEG that have beenused to derivatize other proteins, such as mono (C1-C10) alkoxy- oraryloxy-polyethylene glycol or polyethylene glycol-maleimide. In certainembodiments, the antibody to be pegylated is an aglycosylated antibody.Methods for pegylating proteins are known in the art and can be appliedto the antibodies of the invention. See for example, EP 0 154 316 byNishimura et al. and EP 0 401 384 by Ishikawa et al.

Methods of Engineering Antibodies

As discussed above, the anti-VSIG1, anti-ILDR1, anti-LOC253012,anti-AI216611, anti-C1ORF32, or anti-FXYD3 antibodies having VH and VKsequences disclosed herein can be used to create new anti-VSIG1,anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32, or anti-FXYD3antibodies, respectively, by modifying the VH and/or VL sequences, orthe constant regions attached thereto. Thus, in another aspect of theinvention, the structural features of an anti-VSIG1, anti-ILDR1,anti-LOC253012, anti-AI216611, anti-C1ORF32, or anti-FXYD3 antibody ofthe invention, are used to create structurally related anti-VSIG1,anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32, or anti-FXYD3antibodies that retain at least one functional property of theantibodies of the invention, such as binding to human VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, or FXYD3, respectively. For example, oneor more CDR regions of one VSIG1, ILDR1, LOC253012, AI216611, C1ORF32,or FXYD3 antibody or mutations thereof, can be combined recombinantlywith known framework regions and/or other CDRs to create additional,recombinantly-engineered, anti-VSIG1, anti-ILDR1, anti-LOC253012,anti-AI216611, anti-C1ORF32, or anti-FXYD3 antibodies of the invention,as discussed above. Other types of modifications include those describedin the previous section. The starting material for the engineeringmethod is one or more of the VH and/or VK sequences provided herein, orone or more CDR regions thereof. To create the engineered antibody, itis not necessary to actually prepare (i.e., express as a protein) anantibody having one or more of the VH and/or VK sequences providedherein, or one or more CDR regions thereof. Rather, the informationcontained in the sequences is used as the starting material to create a“second generation” sequences derived from the original sequences andthen the “second generation” sequences is prepared and expressed as aprotein.

Standard molecular biology techniques can be used to prepare and expressaltered antibody sequence.

Preferably, the antibody encoded by the altered antibody sequences isone that retains one, some or all of the functional properties of theanti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32, oranti-FXYD3 antibodies, respectively, produced by methods and withsequences provided herein, which functional properties include bindingto VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3 antigen with aspecific KD level or less and/or modulating B7 costimulation and/orselectively binding to desired target cells such as lung cancer, ovariancancer, colon cancer, that express VSIG1, ILDR1, LOC253012, AI216611,C1ORF32, or FXYD3 antigen.

The functional properties of the altered antibodies can be assessedusing standard assays available in the art and/or described herein.

In certain embodiments of the methods of engineering antibodies of theinvention, mutations can be introduced randomly or selectively along allor part of an anti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611,anti-C1ORF32, or anti-FXYD3 antibody coding sequence and the resultingmodified anti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611,anti-C1ORF32, or anti-FXYD3 antibodies can be screened for bindingactivity and/or other desired functional properties.

Mutational methods have been described in the art. For example, PCTPublication WO 02/092780 by Short describes methods for creating andscreening antibody mutations using saturation mutagenesis, syntheticligation assembly, or a combination thereof. Alternatively, PCTPublication WO 03/074679 by Lazar et al. describes methods of usingcomputational screening methods to optimize physiochemical properties ofantibodies.

Nucleic Acid Molecules Encoding Antibodies of the Invention

Another aspect of the invention pertains to nucleic acid molecules thatencode the antibodies of the invention. The nucleic acids may be presentin whole cells, in a cell lysate, or in a partially purified orsubstantially pure form. A nucleic acid is “isolated” or “renderedsubstantially pure” when purified away from other cellular components orother contaminants, e.g., other cellular nucleic acids or proteins, bystandard techniques, including alkaline/SDS treatment, CsCl banding,column chromatography, agarose gel electrophoresis and others well knownin the art. See, F. Ausubel, et al., ed. (1987) Current Protocols inMolecular Biology, Greene Publishing and Wiley Interscience, New York. Anucleic acid of the invention can be, for example, DNA or RNA and may ormay not contain intronic sequences. In a preferred embodiment, thenucleic acid is a cDNA molecule.

Nucleic acids of the invention can be obtained using standard molecularbiology techniques. For antibodies expressed by hybridomas (e.g.,hybridomas prepared from transgenic mice carrying human immunoglobulingenes as described further below), cDNAs encoding the light and heavychains of the antibody made by the hybridoma can be obtained by standardPCR amplification or cDNA cloning techniques. For antibodies obtainedfrom an immunoglobulin gene library (e.g., using phage displaytechniques), nucleic acid encoding the antibody can be recovered fromthe library.

Once DNA fragments encoding VH and VL segments are obtained, these DNAfragments can be further manipulated by standard recombinant DNAtechniques, for example to convert the variable region genes tofull-length antibody chain genes, to Fab fragment genes or to a scFvgene. In these manipulations, a VL- or VH-encoding DNA fragment isoperatively linked to another DNA fragment encoding another protein,such as an antibody constant region or a flexible linker.

The term “operatively linked”, as used in this context, is intended tomean that the two DNA fragments are joined such that the amino acidsequences encoded by the two DNA fragments remain in-frame.

The isolated DNA encoding the VH region can be converted to afull-length heavy chain gene by operatively linking the VH-encoding DNAto another DNA molecule encoding heavy chain constant regions (CH1, CH2and CH3). The sequences of human heavy chain constant region genes areknown in the art (see e.g., Kabat, E. A., et al. (1991) Sequences ofProteins of Immunological Interest, Fifth Edition, U.S. Department ofHealth and Human Services, NIH Publication No. 91-3242) and DNAfragments encompassing these regions can be obtained by standard PCRamplification. The heavy chain constant region can be an IgG1, IgG2,IgG3, IgG4, IgA, IgE, IgM or IgD constant region, but most preferably isan IgG1 or IgG4 constant region. For a Fab fragment heavy chain gene,the VH-encoding DNA can be operatively linked to another DNA moleculeencoding only the heavy chain CH1 constant region.

The isolated DNA encoding the VL region can be converted to afull-length light chain gene (as well as a Fab light chain gene) byoperatively linking the VL-encoding DNA to another DNA molecule encodingthe light chain constant region, CL. The sequences of human light chainconstant region genes are known in the art (see e.g., Kabat, E. A., etal. (1991) Sequences of Proteins of Immunological Interest, FifthEdition, U.S. Department of Health and Human Services, NIH PublicationNo. 91-3242) and DNA fragments encompassing these regions can beobtained by standard PCR amplification. The light chain constant regioncan be a kappa or lambda constant region, but most preferably is a kappaconstant region.

To create a scFv gene, the VH- and VL-encoding DNA fragments areoperatively linked to another fragment encoding a flexible linker, e.g.,encoding the amino acid sequence (Gly4-Ser)3, such that the VH and VLsequences can be expressed as a contiguous single-chain protein, withthe VL and VH regions joined by the flexible linker (see e.g., Bird etal. (1988) Science 242:423-426; Huston et al. (1988) Proc. Natl. Acad.Sci. USA 85:5879-5883; McCafferty et al., (1990) Nature 348:552-554).

Production of Anti-VSIG1, Anti-ILDR1, Anti-LOC253012, Anti-AI216611,Anti-C1ORF32, or Anti-FXYD3 Monoclonal Antibodies of the Invention

Monoclonal antibodies (mAbs) of the present invention can be produced bya variety of techniques, including conventional monoclonal antibodymethodology e.g., the standard somatic cell hybridization technique ofKohler and Milstein (1975) Nature 256:495. Although somatic cellhybridization procedures are preferred, in principle, other techniquesfor producing monoclonal antibody can be employed e.g., viral oroncogenic transformation of B lymphocytes.

A preferred animal system for preparing hybridomas is the murine system.Hybridoma production in the mouse is a very well-established procedure.Immunization protocols and techniques for isolation of immunizedsplenocytes for fusion are known in the art. Fusion partners (e.g.,murine myeloma cells) and fusion procedures are also known.

Chimeric or humanized antibodies of the present invention can beprepared based on the sequence of a murine monoclonal antibody preparedas described above. DNA encoding the heavy and light chainimmunoglobulins can be obtained from the murine hybridoma of interestand engineered to contain non-murine (e.g., human) immunoglobulinsequences using standard molecular biology techniques. For example, tocreate a chimeric antibody, the murine variable regions can be linked tohuman constant regions using methods known in the art (see e.g., U.S.Pat. No. 4,816,567 to Cabilly et al.). To create a humanized antibody,the murine CDR regions can be inserted into a human framework usingmethods known in the art (see e.g., U.S. Pat. No. 5,225,539 to Winter,and U.S. Pat. Nos. 5,530,101; 5,585,089; 5,693,762 and 6,180,370 toQueen et al.).

In a preferred embodiment, the antibodies of the invention are humanmonoclonal antibodies. Such human monoclonal antibodies directed againstVSIG1 can be generated using transgenic or transchromosomic micecarrying parts of the human immune system rather than the mouse system.These transgenic and transchromosomic mice include mice referred toherein as the HuMAb Mouse® and KM Mouse® respectively, and arecollectively referred to herein as “human Ig mice.” The HuMAb Mouse™(Medarex. Inc.) contains human immunoglobulin gene miniloci that encodeunrearranged human heavy (.mu. and .gamma.) and .kappa. light chainimmunoglobulin sequences, together with targeted mutations thatinactivate the endogenous.mu. and .kappa. chain loci (see e.g., Lonberg,et al. (1994) Nature 368(6474): 856-859). Accordingly, the mice exhibitreduced expression of mouse IgM or .kappa., and in response toimmunization, the introduced human heavy and light chain transgenesundergo class switching and somatic mutation to generate high affinityhuman IgGkappa. monoclonal (Lonberg, N. et al. (1994), supra; reviewedin Lonberg, N. (1994) Handbook of Experimental Pharmacology 113:49-101;Lonberg, N. and Huszar, D. (1995) Intern. Rev. Immunol. 13: 65-93, andHarding, F. and Lonberg, N. (1995) Ann. N.Y. Acad. Sci. 764:536-546).The preparation and use of the HuMab Mouse®, and the genomicmodifications carried by such mice, is further described in Taylor, L.et al. (1992) Nucleic Acids Research 20:6287-6295; Chen, J. et al.(1993) International Immunology 5:647-656; Tuaillon et al. (1993) Proc.Natl. Acad. Sci. USA 90:3720-3724; Choi et al. (1993) Nature Genetics4:117-123; Chen, J. et al. (1993) EMBO J. 12: 821-830; Tuaillon et al.(1994) J. Immunol. 152:2912-2920; Taylor, L. et al. (1994) InternationalImmunology 6:579-591; and Fishwild, D. et al. (1996) NatureBiotechnology 14: 845-851, the contents of all of which are herebyspecifically incorporated by reference in their entirety. See further,U.S. Pat. Nos. 5,545,806; 5,569,825; 5,625,126; 5,633,425; 5,789,650;5,877,397; 5,661,016; 5,814,318; 5,874,299; and 5,770,429; all toLonberg and Kay; U.S. Pat. No. 5,545,807 to Surani et al.; PCTPublication Nos. WO 92/03918, WO 93/12227, WO 94/25585, WO 97/13852, WO98/24884 and WO 99/45962, all to Lonberg and Kay; and PCT PublicationNo. WO 01/14424 to Korman et al.

In another embodiment, human antibodies of the invention can be raisedusing a mouse that carries human immunoglobulin sequences on transgenesand transchromosomes, such as a mouse that carries a human heavy chaintransgene and a human light chain transchromosome. Such mice, referredto herein as “KM Mice™”, are described in detail in PCT Publication WO02/43478 to Ishida et al.

Still further, alternative transgenic animal systems expressing humanimmunoglobulin genes are available in the art and can be used to raiseanti-VSIG1 antibodies of the invention. For example, an alternativetransgenic system referred to as the Xenomouse (Abgenix, Inc.) can beused; such mice are described in, for example, U.S. Pat. Nos. 5,939,598;6,075,181; 6,114,598; 6, 150,584 and 6,162,963 to Kucherlapati et al.

Moreover, alternative transchromosomic animal systems expressing humanimmunoglobulin genes are available in the art and can be used to raiseanti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32, oranti-FXYD3 antibodies of the invention. For example, mice carrying botha human heavy chain transchromosome and a human light chaintranschromosome, referred to as “TC mice” can be used; such mice aredescribed in Tomizuka et al. (2000) Proc. Natl. Acad. Sci. USA97:722-727. Furthermore, cows carrying human heavy and light chaintranschromosomes have been described in the art (Kuroiwa et al. (2002)Nature Biotechnology 20:889-894) and can be used to raise anti-VSIG1antibodies of the invention.

Human monoclonal antibodies of the invention can also be prepared usingphage display methods for screening libraries of human immunoglobulingenes. Such phage display methods for isolating human antibodies areestablished in the art. See for example: U.S. Pat. Nos. 5,223,409;5,403,484; and 5,571,698 to Ladner et al.; U.S. Pat. Nos. 5,427,908 and5,580,717 to Dower et al.; U.S. Pat. Nos. 5,969,108 and 6,172,197 toMcCafferty et al.; and U.S. Pat. Nos. 5,885,793; 6,521,404; 6,544,73 1;6,555,313; 6,582,915 and 6,593,081 to Griffiths et al.

Human monoclonal antibodies of the invention can also be prepared usingSCID mice into which human immune cells have been reconstituted suchthat a human antibody response can be generated upon immunization. Suchmice are described in, for example, U.S. Pat. Nos. 5,476,996 and5,698,767 to Wilson et al.

Immunization of Human Ig Mice

When human Ig mice are used to raise human antibodies of the invention,such mice can be immunized with a purified or enriched preparation ofVSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3 antigen and/orrecombinant VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXY, or anVSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3 fusion protein, asdescribed by Lonberg, N. et al. (1994) Nature 368(6474): 856-859;Fishwild, D. et al. (1996) Nature Biotechnology 14: 845-851; and PCTPublication WO 98/24884 and WO 01/14424. Preferably, the mice will be6-16 weeks of age upon the first infusion. For example, a purified orrecombinant preparation (5-50. mu.g) of VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, or FXYD3 antigen can be used to immunize the human Igmice intraperitoneally.

Prior experience with various antigens by others has shown that thetransgenic mice respond when initially immunized intraperitoneally (IP)with antigen in complete Freund's adjuvant, followed by every other weekIP immunizations (up to a total of 6) with antigen in incompleteFreund's adjuvant. However, adjuvants other than Freund's are also foundto be effective. In addition, whole cells in the absence of adjuvant arefound to be highly immunogenic. The immune response can be monitoredover the course of the immunization protocol with plasma samples beingobtained by retroorbital bleeds. The plasma can be screened by ELISA (asdescribed below), and mice with sufficient titers of anti-VSIG1,anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32, or anti-FXYD3human immunoglobulin can be used for fusions. Mice can be boostedintravenously with antigen 3 days before sacrifice and removal of thespleen. It is expected that 2-3 fusions for each immunization may needto be performed. Between 6 and 24 mice are typically immunized for eachantigen. Usually both HCo7 and HCo12 strains are used. In addition, bothHCo7 and HCo12 transgene can be bred together into a single mouse havingtwo different human heavy chain transgenes (HCo7/HCo 12). Alternativelyor additionally, the KM Mouse® strain can be used.

Generation of Hybridomas Producing Human Monoclonal Antibodies of theInvention

To generate hybridomas producing human monoclonal antibodies of theinvention, splenocytes and/or lymph node cells from immunized mice canbe isolated and fused to an appropriate immortalized cell line, such asa mouse myeloma cell line. The resulting hybridomas can be screened forthe production of antigen-specific antibodies. For example, single cellsuspensions of splenic lymphocytes from immunized mice can be fused toone-sixth the number of P3X63-Ag8.653 nonsecreting mouse myeloma cells(ATCC, CRL 1580) with 50% PEG. Cells are plated at approximately 2×10−5in flat bottom microtiter plate, followed by a two week incubation inselective medium containing 20% fetal Clone Serum, 18% “653” conditionedmedia, 5% origen (IGEN), 4 mM L-glutamine, 1 mM sodium pyruvate, 5 mMHEPES, 0.055 mM 2-mercaptoethanol, 50 units/ml penicillin, 50 mg/mlstreptomycin, 50 mg/ml gentamycin and 1×HAT (Sigma; the HAT is added 24hours after the fusion). After approximately two weeks, cells can becultured in medium in which the HAT is replaced with HT. Individualwells can then be screened by ELISA for human monoclonal IgM and IgGantibodies. Once extensive hybridoma growth occurs, medium can beobserved usually after 10-14 days. The antibody secreting hybridomas canbe replated, screened again, and if still positive for human IgG, themonoclonal antibodies can be subcloned at least twice by limitingdilution. The stable subclones can then be cultured in vitro to generatesmall amounts of antibody in tissue culture medium for characterization.

To purify human monoclonal antibodies, selected hybridomas can be grownin two-liter spinner-flasks for monoclonal antibody purification.Supernatants can be filtered and concentrated before affinitychromatography with protein A-Sepharose (Pharmacia, Piscataway, N.J.).Eluted IgG can be checked by gel electrophoresis and high performanceliquid chromatography to ensure purity. The buffer solution can beexchanged into PBS, and the concentration can be determined by OD280using 1.43 extinction coefficient. The monoclonal antibodies can bealiquoted and stored at −80 degrees C.

Generation of Transfectomas Producing Monoclonal Antibodies of theInvention

Antibodies of the invention also can be produced in a host celltransfectoma using, for example, a combination of recombinant DNAtechniques and gene transfection methods as is well known in the art(e.g., Morrison, S. (1985) Science 229:1202).

For example, to express the antibodies, or antibody fragments thereof,DNAs encoding partial or full-length light and heavy chains, can beobtained by standard molecular biology techniques (e.g., PCRamplification or cDNA cloning using a hybridoma that expresses theantibody of interest) and the DNAs can be inserted into expressionvectors such that the genes are operatively linked to transcriptionaland translational control sequences. In this context, the term“operatively linked” is intended to mean that an antibody gene isligated into a vector such that transcriptional and translationalcontrol sequences within the vector serve their intended function ofregulating the transcription and translation of the antibody gene. Theexpression vector and expression control sequences are chosen to becompatible with the expression host cell used. The antibody light chaingene and the antibody heavy chain gene can be inserted into separatevector or, more typically, both genes are inserted into the sameexpression vector. The antibody genes are inserted into the expressionvector by standard methods (e.g., ligation of complementary restrictionsites on the antibody gene fragment and vector, or blunt end ligation ifno restriction sites are present). The light and heavy chain variableregions of the antibodies described herein can be used to createfull-length antibody genes of any antibody isotype by inserting theminto expression vectors already encoding heavy chain constant and lightchain constant regions of the desired isotype such that the VH segmentis operatively linked to the CH segments within the vector and the VKsegment is operatively linked to the CL segment within the vector.Additionally or alternatively, the recombinant expression vector canencode a signal peptide that facilitates secretion of the antibody chainfrom a host cell. The antibody chain gene can be cloned into the vectorsuch that the signal peptide is linked in-frame to the amino terminus ofthe antibody chain gene. The signal peptide can be an immunoglobulinsignal peptide or a heterologous signal peptide (i.e., a signal peptidefrom a non-immunoglobulin protein).

In addition to the antibody chain genes, the recombinant expressionvectors of the invention carry regulatory sequences that control theexpression of the antibody chain genes in a host cell. The term“regulatory sequence” is intended to include promoters, enhancers andother expression control elements (e.g., polyadenylation signals) thatcontrol the transcription or translation of the antibody chain genes.Such regulatory sequences are described, for example, in Goeddel (GeneExpression Technology. Methods in Enzymology 185, Academic Press, SanDiego, Calif. (1990)). It will be appreciated by those skilled in theart that the design of the expression vector, including the selection ofregulatory sequences, may depend on such factors as the choice of thehost cell to be transformed, the level of expression of protein desired,etc. Preferred regulatory sequences for mammalian host cell expressioninclude viral elements that direct high levels of protein expression inmammalian cells, such as promoters and/or enhancers derived fromcytomegalovirus (CMV), Simian Virus 40 (SV40), adenovirus, (e.g., theadenovirus major late promoter (AdMLP) and polyoma. Alternatively,nonviral regulatory sequences may be used, such as the ubiquitinpromoter or .beta.-globin promoter. Still further, regulatory elementscomposed of sequences from different sources, such as the SR alpha.promoter system, which contains sequences from the SV40 early promoterand the long terminal repeat of human T cell leukemia virus type 1(Takebe, Y. et al. (1988) Mol. Cell. Biol. 8:466-472).

In addition to the antibody chain genes and regulatory sequences, therecombinant expression vectors of the invention may carry additionalsequences, such as sequences that regulate replication of the vector inhost cells (e.g., origins of replication) and selectable marker genes.The selectable marker gene facilitates selection of host cells intowhich the vector has been introduced (see, e.g., U.S. Pat. Nos.4,399,216, 4,634,665 and 5,179,017, all by Axel et al.). For example,typically the selectable marker gene confers resistance to drugs, suchas G418, hygromycin or methotrexate, on a host cell into which thevector has been introduced. Preferred selectable marker genes includethe dihydrofolate reductase (DHFR) gene (for use in dhfr-host cells withmethotrexate selection/amplification) and the neo gene (for G418selection).

For expression of the light and heavy chains, the expression vectorsencoding the heavy and light chains is transfected into a host cell bystandard techniques. The various forms of the term “transfection” areintended to encompass a wide variety of techniques commonly used for theintroduction of exogenous DNA into a prokaryotic or eukaryotic hostcell, e.g., electroporation, calcium-phosphate precipitation,DEAE-dextran transfection and the like. Although it is theoreticallypossible to express the antibodies of the invention in eitherprokaryotic or eukaryotic host cells, expression of antibodies ineukaryotic cells, and most preferably mammalian host cells, is the mostpreferred because such eukaryotic cells, and in particular mammaliancells, are more likely than prokaryotic cells to assemble and secrete aproperly folded and immunologically active antibody. Prokaryoticexpression of antibody genes has been reported to be ineffective forproduction of high yields of active antibody (Boss, M. A. and Wood, C.R. (1985) Immunology Today 6:12-13).

Preferred mammalian host cells for expressing the recombinant antibodiesof the invention include Chinese Hamster Ovary (CHO cells) (includingdhfr-CHO cells, described in Urlaub and Chasin, (1980) Proc. Natl. Acad.Sci. USA 77:4216-4220, used with a DHFR selectable marker, e.g., asdescribed in R. J. Kaufman and P. A. Sharp (1982) Mol. Biol.159:601-621), NSO myeloma cells, COS cells and SP2 cells. In particular,for use with NSO myeloma cells, another preferred expression system isthe GS gene expression system disclosed in WO 87/04462, WO 89/01036 andEP 338,841. When recombinant expression vectors encoding antibody genesare introduced into mammalian host cells, the antibodies are produced byculturing the host cells for a period of time sufficient to allow forexpression of the antibody in the host cells or, more preferably,secretion of the antibody into the culture medium in which the hostcells are grown. Antibodies can be recovered from the culture mediumusing standard protein purification methods.

Characterization of Antibody Binding to Antigen

Antibodies of the invention can be tested for binding to VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, or FXYD3 by, for example, standard ELISA.Briefly, microtiter plates are coated with purified VSIG1 at 0.25.mu.g/ml in PBS, and then blocked with 5% bovine serum albumin in PBS.Dilutions of antibody (e.g., dilutions of plasma from VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, or FXYD3-immunized mice) are added to eachwell and incubated for 1-2 hours at 37 degrees C. The plates are washedwith PBS/Tween and then incubated with secondary reagent (e.g., forhuman antibodies, a goat-anti-human IgG Fc-specific polyclonal reagent)conjugated to alkaline phosphatase for 1 hour at 37 degrees C. Afterwashing, the plates are developed with pNPP substrate (1 mg/ml), andanalyzed at OD of 405-650. Preferably, mice which develop the highesttiters will be used for fusions.

An ELISA assay as described above can also be used to screen forhybridomas that show positive reactivity with VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, or FXYD3 immunogen. Hybridomas that bind with highavidity to VSIG1 are subcloned and further characterized. One clone fromeach hybridoma, which retains the reactivity of the parent cells (byELISA), can be chosen for making a 5-10 vial cell bank stored at −140degrees C., and for antibody purification.

To purify anti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611,anti-C1ORF32, or anti-FXYD3 antibodies, selected hybridomas can be grownin two-liter spinner-flasks for monoclonal antibody purification.Supernatants can be filtered and concentrated before affinitychromatography with protein A-sepharose (Pharmacia, Piscataway, N.J.).Eluted IgG can be checked by gel electrophoresis and high performanceliquid chromatography to ensure purity. The buffer solution can beexchanged into PBS, and the concentration can be determined by OD280using 1.43 extinction coefficient. The monoclonal antibodies can bealiquoted and stored at −80 degrees C.

To determine if the selected anti-VSIG1, anti-ILDR1, anti-LOC253012,anti-AI216611, anti-C1ORF32, or anti-FXYD3 monoclonal antibodies bind tounique epitopes, each antibody can be biotinylated using commerciallyavailable reagents (Pierce, Rockford, Ill.). Competition studies usingunlabeled monoclonal antibodies and biotinylated monoclonal antibodiescan be performed using VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, orFXYD3 coated-ELISA plates as described above. Biotinylated mAb bindingcan be detected with a strepavidin-alkaline phosphatase probe.

To determine the isotype of purified antibodies, isotype ELISAs can beperformed using reagents specific for antibodies of a particularisotype. For example, to determine the isotype of a human monoclonalantibody, wells of microtiter plates can be coated with 1. mu.g/ml ofanti-human immunoglobulin overnight at 4 degrees C. After blocking with1% BSA, the plates are reacted with 1 mug/ml or less of test monoclonalantibodies or purified isotype controls, at ambient temperature for oneto two hours. The wells can then be reacted with either human IgG1 orhuman IgM-specific alkaline phosphatase-conjugated probes. Plates aredeveloped and analyzed as described above.

Anti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32, oranti-FXYD3 human IgGs can be further tested for reactivity with VSIG1,ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3 antigen, respectively, byWestern blotting. Briefly, VSIG1, ILDR1, LOC253012, AI216611, C1ORF32,or FXYD3 antigen can be prepared and subjected to sodium dodecyl sulfatepolyacrylamide gel electrophoresis. After electrophoresis, the separatedantigens are transferred to nitrocellulose membranes, blocked with 10%fetal calf serum, and probed with the monoclonal antibodies to betested. Human IgG binding can be detected using anti-human IgG alkalinephosphatase and developed with BCIP/NBT substrate tablets (Sigma Chem.Co., St. Louis, Mo.).

Conjugates or Immunoconjugates

The present invention encompasses conjugates for use in immune therapycomprising the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3antigen and soluble portions thereof including the ectodomain orportions or variants thereof. For example the invention encompassesconjugates wherein the ECD of the VSIG1, ILDR1, LOC253012, AI216611,C1ORF32, or FXYD3 antigen is attached to an immunoglobulin or fragmentthereof. The invention contemplates the use thereof for promoting orinhibiting VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3 antigenactivities such as immune costimulation and the use thereof in treatingtransplant, autoimmune, and cancer indications described herein.

In another aspect, the present invention features immunoconjugatescomprising an anti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611,anti-C1ORF32, or anti-FXYD3 antibody, or a fragment thereof, conjugatedto a therapeutic moiety, such as a cytotoxin, a drug (e.g., animmunosuppressant) or a radiotoxin. Such conjugates are referred toherein as “immunoconjugates”. Immunoconjugates that include one or morecytotoxins are referred to as “immunotoxins.” A cytotoxin or cytotoxicagent includes any agent that is detrimental to (e.g., kills) cells.Examples include taxol, cytochalasin B, gramicidin D, ethidium bromide,emetine, mitomycin, etoposide, tenoposide, vincristine, vinblastine,colchicin, doxorubicin, daunorubicin, dihydroxy anthracin dione,mitoxantrone, mithramycin, actinomycin D, 1-dehydrotestosterone,glucocorticoids, procaine, tetracaine, lidocaine, propranolol, andpuromycin and analogs or homologs thereof. Therapeutic agents alsoinclude, for example, antimetabolites (e.g., methotrexate,6-mercaptopurine, 6-thioguanine, cytarabine, 5-fluorouracildecarbazine), alkylating agents (e.g., mechlorethamine, thioepachlorambucil, melphalan, carmustine (BSNU) and lomustine (CCNU),cyclothosphamide, busulfan, dibromomannitol, streptozotocin, mitomycinC, and cis-dichlorodiamine platinum (II) (DDP) cisplatin),anthracyclines (e.g., daunorubicin (formerly daunomycin) anddoxorubicin), antibiotics (e.g., dactinomycin (formerly actinomycin),bleomycin, mithramycin, and anthramycin (AMC)), and anti-mitotic agents(e.g., vincristine and vinblastine).

Other preferred examples of therapeutic cytotoxins that can beconjugated to an antibody of the invention include duocarmycins,calicheamicins, maytansines and auristatins, and derivatives thereof. Anexample of a calicheamicin antibody conjugate is commercially available(Mylotarg™; Wyeth).

Cytotoxins can be conjugated to antibodies of the invention using linkertechnology available in the art. Examples of linker types that have beenused to conjugate a cytotoxin to an antibody include, but are notlimited to, hydrazones, thioethers, esters, disulfides andpeptide-containing linkers. A linker can be chosen that is, for example,susceptible to cleavage by low pH within the lysosomal compartment orsusceptible to cleavage by proteases, such as proteases preferentiallyexpressed in tumor tissue such as cathepsins (e.g., cathepsins B, C, D).

For further discussion of types of cytotoxins, linkers and methods forconjugating therapeutic agents to antibodies, see also Saito, G. et al.(2003) Adv. Drug Deliv. Rev. 55:199-215; Trail, P. A. et al. (2003)Cancer Immunol. Immunother. 52:328-337; Payne, G. (2003) Cancer Cell3:207-212; Allen, T. M. (2002) Nat. Rev. Cancer 2:750-763; Pastan, I.and Kreitman, R. J. (2002) Curr. Opin. Investig. Drugs 3:1089-1091;Senter, P. D. and Springer, C. J. (2001) Adv. Drug Deliv. Rev.53:247-264.

Antibodies of the present invention also can be conjugated to aradioactive isotope to generate cytotoxic radiopharmaceuticals, alsoreferred to as radioimmunoconjugates. Examples of radioactive isotopesthat can be conjugated to antibodies for use diagnostically ortherapeutically include, but are not limited to, iodine 131, indium 111,yttrium 90 and lutetium 177. Method for preparing radioimmunconjugatesare established in the art. Examples of radioimmunoconjugates arecommercially available, including Zevalin™ (IDEC Pharmaceuticals) andBexxar™ (Corixa Pharmaceuticals), and similar methods can be used toprepare radioimmunoconjugates using the antibodies of the invention.

The antibody conjugates of the invention can be used to modify a givenbiological response, and the drug moiety is not to be construed aslimited to classical chemical therapeutic agents. For example, the drugmoiety may be a protein or polypeptide possessing a desired biologicalactivity. Such proteins may include, for example, an enzymaticallyactive toxin, or active fragment thereof, such as abrin, ricin A,pseudomonas exotoxin, or diphtheria toxin; a protein such as tumornecrosis factor or interferon-.gamma.; or, biological response modifierssuch as, for example, lymphokines, interleukin-1 (“IL-1”), interleukin-2(“IL-2”), interleukin-6 (“IL-6”), granulocyte macrophage colonystimulating factor (“GM-CSF”), granulocyte colony stimulating factor(“G-CSF”), or other growth factors.

Techniques for conjugating such therapeutic moiety to antibodies arewell known, see, e.g., Arnon et al., “Monoclonal Antibodies ForImmunotargeting Of Drugs In Cancer Therapy”, in Monoclonal AntibodiesAnd Cancer Therapy, Reisfeld et al. (eds.), pp. 243-56 (Alan R. Liss,Inc. 1985); Hellstrom et al., “Antibodies For Drug Delivery”, inControlled Drug Delivery (2nd Ed.), Robinson et al. (eds.), pp. 623-53(Marcel Dekker, Inc. 1987); Thorpe, “Antibody Carriers Of CytotoxicAgents In Cancer Therapy: A Review”, in Monoclonal Antibodies '84:Biological And Clinical Applications, Pinchera et al. (eds.), pp.475-506 (1985); “Analysis, Results, And Future Prospective Of TheTherapeutic Use Of Radiolabeled Antibody In Cancer Therapy”, inMonoclonal Antibodies For Cancer Detection And Therapy, Baldwin et al.(eds.), pp. 303-16 (Academic Press 1985), and Thorpe et al., “ThePreparation And Cytotoxic Properties Of Antibody-Toxin Conjugates”,Immunol. Rev., 62:119-58 (1982).

Bispecific Molecules

In another aspect, the present invention features bispecific moleculescomprising an anti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611,anti-C1ORF32, or anti-FXYD3 antibody, or a fragment thereof, of theinvention. An antibody of the invention, or antigen-binding portionsthereof, can be derivatized or linked to another functional molecule,e.g., another peptide or protein (e.g., another antibody or ligand for areceptor) to generate a bispecific molecule that binds to at least twodifferent binding sites or target molecules. The antibody of theinvention may in fact be derivatized or linked to more than one otherfunctional molecule to generate multispecific molecules that bind tomore than two different binding sites and/or target molecules; suchmultispecific molecules are also intended to be encompassed by the term“bispecific molecule” as used herein. To create a bispecific molecule ofthe invention, an antibody of the invention can be functionally linked(e.g., by chemical coupling, genetic fusion, noncovalent association orotherwise) to one or more other binding molecules, such as anotherantibody, antibody fragment, peptide or binding mimetic, such that abispecific molecule results.

Accordingly, the present invention includes bispecific moleculescomprising at least one first binding specificity for VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, or FXYD3 and a second binding specificityfor a second target epitope. In a particular embodiment of theinvention, the second target epitope is an Fc receptor, e.g., human Fcgamma RI (CD64) or a human Fc alpha receptor (CD89). Therefore, theinvention includes bispecific molecules capable of binding both to Fcgamma. R, Fc alpha R or Fc epsilon R expressing effector cells (e.g.,monocytes, macrophages or polymorphonuclear cells (PMNs)), and to targetcells expressing VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3,respectively. These bispecific molecules target VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, or FXYD3 expressing cells to effector cell andtrigger Fc receptor-mediated effector cell activities, such asphagocytosis of an VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3expressing cells, antibody dependent cell-mediated cytotoxicity (ADCC),cytokine release, or generation of superoxide anion.

In an embodiment of the invention in which the bispecific molecule ismultispecific, the molecule can further include a third bindingspecificity, in addition to an anti-Fc binding specificity and ananti-6f binding specificity. In one embodiment, the third bindingspecificity is an anti-enhancement factor (EF) portion, e.g., a moleculewhich binds to a surface protein involved in cytotoxic activity andthereby increases the immune response against the target cell.

The “anti-enhancement factor portion” can be an antibody, functionalantibody fragment or a ligand that binds to a given molecule, e.g., anantigen or a receptor, and thereby results in an enhancement of theeffect of the binding determinants for the Fc receptor or target cellantigen. The “anti-enhancement factor portion” can bind an Fc receptoror a target cell antigen. Alternatively, the anti-enhancement factorportion can bind to an entity that is different from the entity to whichthe first and second binding specificities bind. For example, theanti-enhancement factor portion can bind a cytotoxic T-cell (e.g., viaCD2, CD3, CD8, CD28, CD4, CD40, ICAM-1 or other immune cell that resultsin an increased immune response against the target cell).

In one embodiment, the bispecific molecules of the invention comprise asa binding specificity at least one antibody, or an antibody fragmentthereof, including, e.g., an Fab, Fab′, F(ab′).sub.2, Fv, or a singlechain Fv. The antibody may also be a light chain or heavy chain dimer,or any minimal fragment thereof such as a Fv or a single chain constructas described in Ladner et al. U.S. Pat. No. 4,946,778, the contents ofwhich is expressly incorporated by reference.

In one embodiment, the binding specificity for an Fcy receptor isprovided by a monoclonal antibody, the binding of which is not blockedby human immunoglobulin G (IgG). As used herein, the term “IgG receptor”refers to any of the eight.gamma.-chain genes located on chromosome 1.These genes encode a total of twelve transmembrane or soluble receptorisoforms which are grouped into three Fc.gamma. receptor classes: Fcgamma R1 (CD64), Fc gamma RII (CD32), and Fc gamma.RIII (CD 16). In onepreferred embodiment, the Fc gamma. receptor a human high affinityFc.gamma R1. The human Fc gammaRI is a 72 kDa molecule, which shows highaffinity for monomeric IgG (10 8-10−9 M.-1).

The production and characterization of certain preferred anti-Fc gamma.monoclonal antibodies are described by Fanger et al. in PCT PublicationWO 88/00052 and in U.S. Pat. No. 4,954,617, the teachings of which arefully incorporated by reference herein. These antibodies bind to anepitope of Fc.gamma.RI, FcyRII or FcyRIII at a site which is distinctfrom the Fc.gamma. binding site of the receptor and, thus, their bindingis not blocked substantially by physiological levels of IgG. Specificanti-Fc.gamma.RI antibodies useful in this invention are mAb 22, mAb 32,mAb 44, mAb 62 and mAb 197. The hybridoma producing mAb 32 is availablefrom the American Type Culture Collection, ATCC Accession No. HB9469. Inother embodiments, the anti-Fcy receptor antibody is a humanized form ofmonoclonal antibody 22 (H22). The production and characterization of theH22 antibody is described in Graziano, R. F. et al. (1995) J. Immunol.155 (10): 4996-5002 and PCT Publication WO 94/10332. The H22 antibodyproducing cell line is deposited at the American Type Culture Collectionunder the designation HAO22CLI and has the accession no. CRL 11177.

In still other preferred embodiments, the binding specificity for an Fcreceptor is provided by an antibody that binds to a human IgA receptor,e.g., an Fc-alpha receptor (Fc alpha.RI (CD89)), the binding of which ispreferably not blocked by human immunoglobulin A (IgA). The term “IgAreceptor” is intended to include the gene product of one alpha.-gene (Fcalpha.RI) located on chromosome 19. This gene is known to encode severalalternatively spliced transmembrane isoforms of 55 to 10 kDa

Fc.alpha.RI (CD89) is constitutively expressed on monocytes/macrophages,eosinophilic and neutrophilic granulocytes, but not on non-effector cellpopulations. Fc alpha R1 has medium affinity (Approximately 5×10−7 M-1)for both IgA1 and IgA2, which is increased upon exposure to cytokinessuch as G-CSF or GM-CSF (Morton, H. C. et al. (1996) Critical Reviews inImmunology 16:423-440). Four FcaRI-specific monoclonal antibodies,identified as A3, A59, A62 and A77, which bind Fc.alpha.RI outside theIgA ligand binding domain, have been described (Monteiro, R. C. et al.(1992) J. Immunol. 148:1764).

Fc. alpha. RI and Fc gamma. R1 are preferred trigger receptors for usein the bispecific molecules of the invention because they are (1)expressed primarily on immune effector cells, e.g., monocytes, PMNs,macrophages and dendritic cells; (2) expressed at high levels (e.g.,5,000-100,000 per cell); (3) mediators of cytotoxic activities (e.g.,ADCC, phagocytosis); (4) mediate enhanced antigen presentation ofantigens, including self-antigens, targeted to them.

While human monoclonal antibodies are preferred, other antibodies whichcan be employed in the bispecific molecules of the invention are murine,chimeric and humanized monoclonal antibodies.

The bispecific molecules of the present invention can be prepared byconjugating the constituent binding specificities, e.g., the anti-FcRand anti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32,or anti-FXYD3 binding specificities, using methods known in the art. Forexample, each binding specificity of the bispecific molecule can begenerated separately and then conjugated to one another. When thebinding specificities are proteins or peptides, a variety of coupling orcross-linking agents can be used for covalent conjugation. Examples ofcross-linking agents include protein A, carbodiimide,N-succinimidyl-5-acetyl-thioacetate (SATA),5,5′-dithiobis(2-nitrobenzoic acid) (DTNB), o-phenylenedimaleimide(oPDM), N-succinimidyl-3-(2-pyridyld-ithio)propionate (SPDP), andsulfosuccinimidyl 4-(N-maleimidomethyl) cyclohaxane-1-carboxylate(sulfo-SMCC) (see e.g., Karpovsky et al. (1984) J. Exp. Med. 160:1686;Liu, M A et al. (1985) Proc. Natl. Acad. Sci. USA 82:8648). Othermethods include those described in Paulus (1985) Behring Ins. Mitt. No.78, 118-132; Brennan et al. (1985) Science 229:81-83), and Glennie etal. (1987) J. Immunol. 139: 2367-2375). Preferred conjugating agents areSATA and sulfo-SMCC, both available from Pierce Chemical Co. (Rockford,Ill.).

When the binding specificities are antibodies, they can be conjugatedvia sulfhydryl bonding of the C-terminus hinge regions of the two heavychains. In a particularly preferred embodiment, the hinge region ismodified to contain an odd number of sulfhydryl residues, preferablyone, prior to conjugation.

Alternatively, both binding specificities can be encoded in the samevector and expressed and assembled in the same host cell. This method isparticularly useful where the bispecific molecule is a mAbXmAb, mAbXFab,FabXF(ab′)2 or ligandXFab fusion protein. A bispecific molecule of theinvention can be a single chain molecule comprising one single chainantibody and a binding determinant, or a single chain bispecificmolecule comprising two binding determinants. Bispecific molecules maycomprise at least two single chain molecules. Methods for preparingbispecific molecules are described for example in U.S. Pat. No.5,260,203; U.S. Pat. No. 5,455,030; U.S. Pat. No. 4,881,175; U.S. Pat.No. 5,132,405; U.S. Pat. No. 5,091,513; U.S. Pat. No. 5,476,786; U.S.Pat. No. 5,013,653; U.S. Pat. No. 5,258,498; and U.S. Pat. No.5,482,858.

Binding of the bispecific molecules to their specific targets can beconfirmed by, for example, enzyme-linked immunosorbent assay (ELISA),radioimmunoassay (RIA), FACS analysis, bioassay (e.g., growthinhibition), or Western Blot assay. Each of these assays generallydetects the presence of protein-antibody complexes of particularinterest by employing a labeled reagent (e.g., an antibody) specific forthe complex of interest. For example, the FcR-antibody complexes can bedetected using e.g., an enzyme-linked antibody or antibody fragmentwhich recognizes and specifically binds to the antibody-FcR complexes.Alternatively, the complexes can be detected using any of a variety ofother immunoassays. For example, the antibody can be radioactivelylabeled and used in a radioimmunoassay (RIA) (see, for example,Weintraub, B., Principles of Radioimmunoassays, Seventh Training Courseon Radioligand Assay Techniques, The Endocrine Society, March, 1986,which is incorporated by reference herein). The radioactive isotope canbe detected by such means as the use of a gamma. counter or ascintillation counter or by autoradiography.

Pharmaceutical Compositions

In another aspect, the present invention provides a composition, e.g., apharmaceutical composition, containing one or a combination ofmonoclonal antibodies, or antigen-binding portions thereof, of thepresent invention, formulated together with a pharmaceuticallyacceptable carrier. Such compositions may include one or a combinationof (e.g., two or more different) antibodies, or immunoconjugates orbispecific molecules of the invention. For example, a pharmaceuticalcomposition of the invention can comprise a combination of antibodies(or immunoconjugates or bispecifics) that bind to different epitopes onthe target antigen or that have complementary activities.

As discussed supra, VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3the invention further embraces identifying other molecules such as smallorganic molecules, peptides, ribozymes, carbohydrates, glycoprotein,siRNAs, antisense RNAs and the like which specifically bind and/ormodulate (enhance or inhibit) an activity elicited by the VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, or FXYD3 antigen, respectively. Thesemolecules may be identified by known screening methods such as bindingassays. Typically these assays will be high throughput and will screen alarge library of synthesized or native compounds in order to identifyputative drug candidates that bind and/or modulate VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, or FXYD3 related activities.

Specifically, the invention embraces the development of drugs containingthe ectodomain of the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, orFXYD3 antigen or a fragment or variant thereof or a correspondingnucleic acid sequence encoding. These conjugates may contain a targetingor other moiety such as an immunoglobulin domain. These conjugates maybe expressed in known vector systems or cells or vectors containing thecorresponding nucleic acid sequences may be used for cancer treatmentand in immune therapy such as in the treatment of autoimmunity,transplant, GVHD, cancer, and other immune disorders or conditions.

Thus, the present invention features a pharmaceutical compositioncomprising a therapeutically effective amount of a therapeutic agentaccording to the present invention. According to the present inventionthe therapeutic agent could be any one of VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, or FXYD3 ectodomain, or a fragment or variantthereof, or a corresponding nucleic acid sequence encoding.

The pharmaceutical composition according to the present invention isfurther preferably used for the treatment of cancers including by way ofexample non-solid and solid tumors, sarcomas, hematological malignanciesincluding but not limited to acute lymphocytic leukemia, chroniclymphocytic leukemia, acute myelogenous leukemia, chronic myelogenousleukemia, multiple myeloma, Hodgkin's lymphoma, Non-Hodgkin's lymphoma,cancer of the breast, prostate, lung, ovary, colon, spleen, kidney,bladder, head and neck, uterus, testicles, stomach, cervix, liver, bone,skin, pancreas, brain and wherein the cancer may be non-metastatic,invasive or metastatic.

The pharmaceutical composition according to the present invention isfurther used for the treatment of autoimmunity and preferably fortreating an autoimmune disease selected from: Multiple sclerosis;Psoriasis; Rheumatoid arthritis; Systemic lupus erythematosus;Ulcerative colitis; Crohn's disease; immune disorders associated withgraft transplantation rejection, benign lymphocytic angiitis, lupuserythematosus, Hashimoto's thyroiditis, primary myxedema, Graves'disease, pernicious anemia, autoimmune atrophic gastritis, Addison'sdisease, insulin dependent diabetes mellitis, good pasture's syndrome,myasthenia gravis, pemphigus, sympathetic ophthalmia, autoimmuneuveitis, autoimmune hemolytic anemia, idiopathic thrombocytopenia,primary biliary cirrhosis, chronic action hepatitis, ulceratis colitis,Sjogren's syndrome, rheumatic disease, polymyositis, scleroderma, mixedconnective tissue disease, inflammatory rheumatism, degenerativerheumatism, extra-articular rheumatism, collagen diseases, chronicpolyarthritis, psoriasis arthropathica, ankylosing spondylitis, juvenilerheumatoid arthritis, periarthritis humeroscapularis, panarteriitisnodosa, progressive systemic scleroderma, arthritis uratica,dermatomyositis, muscular rheumatism, myositis, myogelosis andchondrocalcinosis.

The pharmaceutical composition according to the present invention ispreferably used for the treatment of for rejection of any organtransplant and/or Graft versus host disease which might develop afterbone marrow transplantation.

“Treatment” refers to both therapeutic treatment and prophylactic orpreventative measures. Those in need of treatment include those alreadywith the disorder as well as those in which the disorder is to beprevented. Hence, the mammal to be treated herein may have beendiagnosed as having the disorder or may be predisposed or susceptible tothe disorder. “Mammal” for purposes of treatment refers to any animalclassified as a mammal, including humans, domestic and farm animals, andzoo, sports, or pet animals, such as dogs, horses, cats, cows, etc.Preferably, the mammal is human.

The term “therapeutically effective amount” refers to an amount of agentaccording to the present invention that is effective to treat a diseaseor disorder in a mammal.

The therapeutic agents of the present invention can be provided to thesubject alone, or as part of a pharmaceutical composition where they aremixed with a pharmaceutically acceptable carrier.

Pharmaceutical compositions of the invention also can be administered incombination therapy, i.e., combined with other agents. For example, thecombination therapy can include an anti-VSIG1, anti-ILDR1,anti-LOC253012, anti-AI216611, anti-C1ORF32, or anti-FXYD3 antibody orVSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3 modulating agentaccording to the present invention such as a soluble polypeptideconjugate containing the ectodomain of the VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, or FXYD3 antigen or a small molecule such as apeptide, ribozyme, siRNA, or other drug that binds VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, or FXYD3 combined with at least one othertherapeutic or immune modulatory agent. Examples of therapeutic agentsthat can be used in combination therapy are described in greater detailbelow in the section on uses of the antibodies of the invention.

As used herein, “pharmaceutically acceptable carrier” includes any andall solvents, dispersion media, coatings, antibacterial and antifungalagents, isotonic and absorption delaying agents, and the like that arephysiologically compatible. Preferably, the carrier is suitable forintravenous, intramuscular, subcutaneous, parenteral, spinal orepidermal administration (e.g., by injection or infusion). Depending onthe route of administration, the active compound, i.e., antibody,immunoconjugate, or bispecific molecule, may be coated in a material toprotect the compound from the action of acids and other naturalconditions that may inactivate the compound. The pharmaceuticalcompounds of the invention may include one or more pharmaceuticallyacceptable salts. A “pharmaceutically acceptable salt” refers to a saltthat retains the desired biological activity of the parent compound anddoes not impart any undesired toxicological effects (see e.g., Berge, S.M., et al. (1977) J. Pharm. Sci. 66: 1-19). Examples of such saltsinclude acid addition salts and base addition salts. Acid addition saltsinclude those derived from nontoxic inorganic acids, such ashydrochloric, nitric, phosphoric, sulfuric, hydrobromic, hydroiodic,phosphorous and the like, as well as from nontoxic organic acids such asaliphatic mono- and dicarboxylic acids, phenyl-substituted alkanoicacids, hydroxy alkanoic acids, aromatic acids, aliphatic and aromaticsulfonic acids and the like. Base addition salts include those derivedfrom alkaline earth metals, such as sodium, potassium, magnesium,calcium and the like, as well as from nontoxic organic amines, such asN,N′-dibenzylethylenediamine, N-methylglucamine, chloroprocaine,choline, diethanolamine, ethylenediamine, procaine and the like.

A pharmaceutical composition of the invention also may include apharmaceutically acceptable anti-oxidant. Examples of pharmaceuticallyacceptable antioxidants include: (1) water soluble antioxidants, such asascorbic acid, cysteine hydrochloride, sodium bisulfate, sodiummetabisulfite, sodium sulfite and the like; (2) oil-solubleantioxidants, such as ascorbyl palmitate, butylated hydroxyanisole(BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate,alpha-tocopherol, and the like; and (3) metal chelating agents, such ascitric acid, ethylenediamine tetraacetic acid (EDTA), sorbitol, tartaricacid, phosphoric acid, and the like.

A pharmaceutical composition of the invention also may include apharmaceutically acceptable anti-oxidant. Examples of pharmaceuticallyacceptable antioxidants include: (1) water soluble antioxidants, such asascorbic acid, cysteine hydrochloride, sodium bisulfate, sodiummetabisulfite, sodium sulfite and the like; (2) oil-solubleantioxidants, such as ascorbyl palmitate, butylated hydroxyanisole(BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate,alpha-tocopherol, and the like; and (3) metal chelating agents, such ascitric acid, ethylenediamine tetraacetic acid (EDTA), sorbitol, tartaricacid, phosphoric acid, and the like. Examples of suitable aqueous andnonaqueous carriers that may be employed in the pharmaceuticalcompositions of the invention include water, ethanol, polyols (such asglycerol, propylene glycol, polyethylene glycol, and the like), andsuitable mixtures thereof, vegetable oils, such as olive oil, andinjectable organic esters, such as ethyl oleate. Proper fluidity can bemaintained, for example, by the use of coating materials, such aslecithin, by the maintenance of the required particle size in the caseof dispersions, and by the use of surfactants.

These compositions may also contain adjuvants such as preservatives,wetting agents, emulsifying agents and dispersing agents. Prevention ofpresence of microorganisms may be ensured both by sterilizationprocedures, supra, and by the inclusion of various antibacterial andantifungal agents, for example, paraben, chlorobutanol, phenol sorbicacid, and the like. It may also be desirable to include isotonic agents,such as sugars, sodium chloride, and the like into the compositions. Inaddition, prolonged absorption of the injectable pharmaceutical form maybe brought about by the inclusion of agents which delay absorption suchas aluminum monostearate and gelatin.

Pharmaceutically acceptable carriers include sterile aqueous solutionsor dispersions and sterile powders for the extemporaneous preparation ofsterile injectable solutions or dispersion. The use of such media andagents for pharmaceutically active substances is known in the art.Except insofar as any conventional media or agent is incompatible withthe active compound, use thereof in the pharmaceutical compositions ofthe invention is contemplated. Supplementary active compounds can alsobe incorporated into the compositions.

Therapeutic compositions typically must be sterile and stable under theconditions of manufacture and storage. The composition can be formulatedas a solution, microemulsion, liposome, or other ordered structuresuitable to high drug concentration. The carrier can be a solvent ordispersion medium containing, for example, water, ethanol, polyol (forexample, glycerol, propylene glycol, and liquid polyethylene glycol, andthe like), and suitable mixtures thereof. The proper fluidity can bemaintained, for example, by the use of a coating such as lecithin, bythe maintenance of the required particle size in the case of dispersionand by the use of surfactants. In many cases, it will be preferable toinclude isotonic agents, for example, sugars, polyalcohols such asmannitol, sorbitol, or sodium chloride in the composition. Prolongedabsorption of the injectable compositions can be brought about byincluding in the composition an agent that delays absorption, forexample, monostearate salts and gelatin. Sterile injectable solutionscan be prepared by incorporating the active compound in the requiredamount in an appropriate solvent with one or a combination ofingredients enumerated above, as required, followed by sterilizationmicrofiltration. Generally, dispersions are prepared by incorporatingthe active compound into a sterile vehicle that contains a basicdispersion medium and the required other ingredients from thoseenumerated above. In the case of sterile powders for the preparation ofsterile injectable solutions, the preferred methods of preparation arevacuum drying and freeze-drying (lyophilization) that yield a powder ofthe active ingredient plus any additional desired ingredient from apreviously sterile-filtered solution thereof.

Sterile injectable solutions can be prepared by incorporating the activecompound in the required amount in an appropriate solvent with one or acombination of ingredients enumerated above, as required, followed bysterilization microfiltration. Generally, dispersions are prepared byincorporating the active compound into a sterile vehicle that contains abasic dispersion medium and the required other ingredients from thoseenumerated above. In the case of sterile powders for the preparation ofsterile injectable solutions, the preferred methods of preparation arevacuum drying and freeze-drying (lyophilization) that yield a powder ofthe active ingredient plus any additional desired ingredient from apreviously sterile-filtered solution thereof.

The amount of active ingredient which can be combined with a carriermaterial to produce a single dosage form will vary depending upon thesubject being treated, and the particular mode of administration. Theamount of active ingredient which can be combined with a carriermaterial to produce a single dosage form will generally be that amountof the composition which produces a therapeutic effect. Generally, outof one hundred percent, this amount will range from about 0.01 percentto about ninety-nine percent of active ingredient, preferably from about0.1 percent to about 70 percent, most preferably from about 1 percent toabout 30 percent of active ingredient in combination with apharmaceutically acceptable carrier.

Dosage regimens are adjusted to provide the optimum desired response(e.g., a therapeutic response). For example, a single bolus may beadministered, several divided doses may be administered over time or thedose may be proportionally reduced or increased as indicated by theexigencies of the therapeutic situation. It is especially advantageousto formulate parenteral compositions in dosage unit form for ease ofadministration and uniformity of dosage. Dosage unit form as used hereinrefers to physically discrete units suited as unitary dosages for thesubjects to be treated; each unit contains a predetermined quantity ofactive compound calculated to produce the desired therapeutic effect inassociation with the required pharmaceutical carrier. The specificationfor the dosage unit forms of the invention are dictated by and directlydependent on (a) the unique characteristics of the active compound andthe particular therapeutic effect to be achieved, and (b) thelimitations inherent in the art of compounding such an active compoundfor the treatment of sensitivity in individuals.

For administration of the antibody, the dosage ranges from about 0.0001to 100 mg/kg, and more usually 0.01 to 5 mg/kg, of the host body weight.For example dosages can be 0.3 mg/kg body weight, 1 mg/kg body weight, 3mg/kg body weight, 5 mg/kg body weight or 10 mg/kg body weight or withinthe range of 1-10 mg/kg. An exemplary treatment regime entailsadministration once per week, once every two weeks, once every threeweeks, once every four weeks, once a month, once every 3 months or onceevery three to 6 months. Preferred dosage regimens for an anti-VSIG1antibody of the invention include 1 mg/kg body weight or 3 mg/kg bodyweight via intravenous administration, with the antibody being givenusing one of the following dosing schedules: (i) every four weeks forsix dosages, then every three months; (ii) every three weeks; (iii) 3mg/kg body weight once followed by 1 mg/kg body weight every threeweeks.

In some methods, two or more monoclonal antibodies with differentbinding specificities are administered simultaneously, in which case thedosage of each antibody administered falls within the ranges indicated.Antibody is usually administered on multiple occasions. Intervalsbetween single dosages can be, for example, weekly, monthly, every threemonths or yearly. Intervals can also be irregular as indicated bymeasuring blood levels of antibody to the target antigen in the patient.In some methods, dosage is adjusted to achieve a plasma antibodyconcentration of about 1-1000 mug/ml and in some methods about 25-300.mu.g/ml.

Alternatively, antibody can be administered as a sustained releaseformulation, in which case less frequent administration is required.Dosage and frequency vary depending on the half-life of the antibody inthe patient. In general, human antibodies show the longest half life,followed by humanized antibodies, chimeric antibodies, and nonhumanantibodies. The dosage and frequency of administration can varydepending on whether the treatment is prophylactic or therapeutic. Inprophylactic applications, a relatively low dosage is administered atrelatively infrequent intervals over a long period of time. Somepatients continue to receive treatment for the rest of their lives. Intherapeutic applications, a relatively high dosage at relatively shortintervals is sometimes required until progression of the disease isreduced or terminated, and preferably until the patient shows partial orcomplete amelioration of symptoms of disease. Thereafter, the patientcan be administered a prophylactic regime.

Actual dosage levels of the active ingredients in the pharmaceuticalcompositions of the present invention may be varied so as to obtain anamount of the active ingredient which is effective to achieve thedesired therapeutic response for a particular patient, composition, andmode of administration, without being toxic to the patient. The selecteddosage level will depend upon a variety of pharmacokinetic factorsincluding the activity of the particular compositions of the presentinvention employed, or the ester, salt or amide thereof, the route ofadministration, the time of administration, the rate of excretion of theparticular compound being employed, the duration of the treatment, otherdrugs, compounds and/or materials used in combination with theparticular compositions employed, the age, sex, weight, condition,general health and prior medical history of the patient being treated,and like factors well known in the medical arts.

A “therapeutically effective dosage” of an anti-VSIG1, anti-ILDR1,anti-LOC253012, anti-AI216611, anti-C1ORF32, or anti-FXYD3 antibody ofthe invention preferably results in a decrease in severity of diseasesymptoms, an increase in frequency and duration of disease symptom-freeperiods, an increase in lifespan, disease remission, or a prevention ofimpairment or disability due to the disease affliction. For example, forthe treatment of VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3positive tumors, e.g., lung tumors, ovarian tumors, and colon tumors, a“therapeutically effective dosage” preferably inhibits cell growth ortumor growth by at least about 20%, more preferably by at least about40%, even more preferably by at least about 60%, and still morepreferably by at least about 80% relative to untreated subjects. Theability of a compound to inhibit tumor growth can be evaluated in ananimal model system predictive of efficacy in human tumors.Alternatively, this property of a composition can be evaluated byexamining the ability of the compound to inhibit, such inhibition invitro by assays known to the skilled practitioner. A therapeuticallyeffective amount of a therapeutic compound can decrease tumor size, orotherwise ameliorate symptoms in a subject. One of ordinary skill in theart would be able to determine such amounts based on such factors as thesubject's size, the severity of the subject's symptoms, and theparticular composition or route of administration selected.

A composition of the present invention can be administered via one ormore routes of administration using one or more of a variety of methodsknown in the art. As will be appreciated by the skilled artisan, theroute and/or mode of administration will vary depending upon the desiredresults. Preferred routes of administration for antibodies of theinvention include intravenous, intramuscular, intradermal,intraperitoneal, subcutaneous, spinal or other parenteral routes ofadministration, for example by injection or infusion. The phrase“parenteral administration” as used herein means modes of administrationother than enteral and topical administration, usually by injection, andincludes, without limitation, intravenous, intramuscular, intraarterial,intrathecal, intracapsular, intraorbital, intracardiac, intradermal,intraperitoneal, transtracheal, subcutaneous, subcuticular,intraarticular, subcapsular, subarachnoid, intraspinal, epidural andintrasternal injection and infusion.

Alternatively, an antibody or other VSIG1, ILDR1, LOC253012, AI216611,C1ORF32, or FXYD3 drug or molecule and their conjugates and combinationsthereof that modulates a VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, orFXYD3 antigen activity according to the invention can be administeredvia a non-parenteral route, such as a topical, epidermal or mucosalroute of administration, for example, intranasally, orally, vaginally,rectally, sublingually or topically.

The active compounds can be prepared with carriers that will protect thecompound against rapid release, such as a controlled releaseformulation, including implants, transdermal patches, andmicroencapsulated delivery systems. Biodegradable, biocompatiblepolymers can be used, such as ethylene vinyl acetate, polyanhydrides,polyglycolic acid, collagen, polyorthoesters, and polylactic acid. Manymethods for the preparation of such formulations are patented orgenerally known to those skilled in the art. See, e.g., Sustained andControlled Release Drug Delivery Systems, J. R. Robinson, ed., MarcelDekker, Inc., New York, 1978.

Therapeutic compositions can be administered with medical devices knownin the art. For example, in a preferred embodiment, a therapeuticcomposition of the invention can be administered with a needleshypodermic injection device, such as the devices disclosed in U.S. Pat.No. 5,399,163; 5,383,851; 5,312,335; 5,064,413; 4,941,880; 4,790,824; or4,596,556. Examples of well-known implants and modules useful in thepresent invention include: U.S. Pat. No. 4,487,603, which discloses animplantable micro-infusion pump for dispensing medication at acontrolled rate; U.S. Pat. No. 4,486,194, which discloses a therapeuticdevice for administering medicaments through the skin; U.S. Pat. No.4,447,233, which discloses a medication infusion pump for deliveringmedication at a precise infusion rate; U.S. Pat. No. 4,447,224, whichdiscloses a variable flow implantable infusion apparatus for continuousdrug delivery; U.S. Pat. No. 4,439,196, which discloses an osmotic drugdelivery system having multi-chamber compartments; and U.S. Pat. No.4,475,196, which discloses an osmotic drug delivery system. Thesepatents are incorporated herein by reference. Many other such implants,delivery systems, and modules are known to those skilled in the art.

In certain embodiments, the antibodies or other VSIG1 related drugs ofthe invention can be formulated to ensure proper distribution in vivo.For example, the blood-brain barrier (BBB) excludes many highlyhydrophilic compounds. To ensure that the therapeutic compounds of theinvention cross the BBB (if desired), they can be formulated, forexample, in liposomes. For methods of manufacturing liposomes, see,e.g., U.S. Pat. Nos. 4,522,811; 5,374,548; and 5,399,331. The liposomesmay comprise one or more moieties which are selectively transported intospecific cells or organs, thus enhance targeted drug delivery (see,e.g., V. V. Ranade (1989) J. Clin. Pharmacol. 29:685). Exemplarytargeting moieties include folate or biotin (see, e.g., U.S. Pat. No.5,416,016 to Low et al.); mannosides (Umezawa et al., (1988) Biochem.Biophys. Res. Commun. 153:1038); antibodies (P. G. Bloeman et al. (1995)FEBS Lett. 357:140; M. Owais et al. (1995) Antimicrob. Agents Chemother.39:180); surfactant protein A receptor (Briscoe et al. (1995) Am. J.Physiol. 1233:134); p120 (Schreier et al. (1994) J. Biol. Chem.269:9090); see also K. Keinanen; M. L. Laukkanen (1994) FEBS Lett.346:123; J. J. Killion; I. J. Fidler (1994) Immunomethods 4:273.

Diagnostic Uses of VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3Antigen and Corresponding Polynucleotides

According to some embodiments, the sample taken from a subject (patient)to perform the diagnostic assay according to the present invention isselected from the group consisting of a body fluid or secretionincluding but not limited to blood, serum, urine, plasma, prostaticfluid, seminal fluid, semen, the external secretions of the skin,respiratory, intestinal, and genitourinary tracts, tears, cerebrospinalfluid, sputum, saliva, milk, peritoneal fluid, pleural fluid, cystfluid, secretions of the breast ductal system (and/or lavage thereof),broncho alveolar lavage, lavage of the reproductive system and lavage ofany other part of the body or system in the body; samples of any organincluding isolated cells or tissues, wherein the cell or tissue can beobtained from an organ selected from, but not limited to lung, colon,ovarian and/or breast tissue; stool or a tissue sample, or anycombination thereof. In some embodiments, the term encompasses samplesof in vivo cell culture constituents. Prior to be subjected to thediagnostic assay, the sample can optionally be diluted with a suitableeluant.

In some embodiments, the phrase “marker” in the context of the presentinvention refers to a nucleic acid fragment, a peptide, or apolypeptide, which is differentially present in a sample taken frompatients (subjects) having one of the herein-described diseases orconditions, as compared to a comparable sample taken from subjects whodo not have one the above-described diseases or conditions.

In some embodiments, the term “polypeptide” is to be understood to referto a molecule comprising from at least 2 to several thousand or moreamino acids. The term “polypeptide” is to be understood to include,inter alia, native peptides (either degradation products, syntheticallysynthesized peptides or recombinant peptides), peptidomimetics, such aspeptoids and semipeptoids or peptide analogs, which may comprise, forexample, any desirable modification, including, inter alia,modifications rendering the peptides more stable while in a body or morecapable of penetrating into cells, or others as will be appreciated byone skilled in the art. Such modifications include, but are not limitedto N terminus modification, C terminus modification, peptide bondmodification, backbone modifications, residue modification, or others.Inclusion of such peptides within the polypeptides of this invention mayproduce a polypeptide sharing identity with the polypeptides describedherein, for example, those provided in the sequence listing.

In some embodiments, the phrase “differentially present” refers todifferences in the quantity or quality of a marker present in a sampletaken from patients having one of the herein-described diseases orconditions as compared to a comparable sample taken from patients who donot have one of the herein-described diseases or conditions. Forexample, a nucleic acid fragment may optionally be differentiallypresent between the two samples if the amount of the nucleic acidfragment in one sample is significantly different from the amount of thenucleic acid fragment in the other sample, for example as measured byhybridization and/or NAT-based assays. A polypeptide is differentiallypresent between the two samples if the amount of the polypeptide in onesample is significantly different from the amount of the polypeptide inthe other sample. It should be noted that if the marker is detectable inone sample and not detectable in the other, then such a marker can beconsidered to be differentially present. Optionally, a relatively lowamount of up-regulation may serve as the marker, as described herein.One of ordinary skill in the art could easily determine such relativelevels of the markers; further guidance is provided in the descriptionof each individual marker below.

In some embodiments, the phrase “diagnostic” means identifying thepresence or nature of a pathologic condition. Diagnostic methods differin their sensitivity and specificity. The “sensitivity” of a diagnosticassay is the percentage of diseased individuals who test positive(percent of “true positives”). Diseased individuals not detected by theassay are “false negatives.” Subjects who are not diseased and who testnegative in the assay are termed “true negatives.” The “specificity” ofa diagnostic assay is 1 minus the false positive rate, where the “falsepositive” rate is defined as the proportion of those without the diseasewho test positive. While a particular diagnostic method may not providea definitive diagnosis of a condition, it suffices if the methodprovides a positive indication that aids in diagnosis.

In some embodiments, the phrase “qualitative” when in reference todifferences in expression levels of a polynucleotide or polypeptide asdescribed herein, refers to the presence versus absence of expression,or in some embodiments, the temporal regulation of expression, or insome embodiments, the timing of expression, or in some embodiments, anypost-translational modifications to the expressed molecule, and others,as will be appreciated by one skilled in the art. In some embodiments,the phrase “quantitative” when in reference to differences in expressionlevels of a polynucleotide or polypeptide as described herein, refers toabsolute differences in quantity of expression, as determined by anymeans, known in the art, or in other embodiments, relative differences,which may be statistically significant, or in some embodiments, whenviewed as a whole or over a prolonged period of time, etc., indicate atrend in terms of differences in expression.

In some embodiments, the term “diagnosing” refers to classifying adisease or a symptom, determining a severity of the disease, monitoringdisease progression, forecasting an outcome of a disease and/orprospects of recovery. The term “detecting” may also optionallyencompass any of the above.

Diagnosis of a disease according to the present invention can, in someembodiments, be affected by determining a level of a polynucleotide or apolypeptide of the present invention in a biological sample obtainedfrom the subject, wherein the level determined can be correlated withpredisposition to, or presence or absence of the disease. It should benoted that a “biological sample obtained from the subject” may alsooptionally comprise a sample that has not been physically removed fromthe subject, as described in greater detail below.

In some embodiments, the term “level” refers to expression levels of RNAand/or protein or to DNA copy number of a marker of the presentinvention.

Typically the level of the marker in a biological sample obtained fromthe subject is different (i.e., increased or decreased) from the levelof the same marker in a similar sample obtained from a healthyindividual (examples of biological samples are described herein).

Numerous well known tissue or fluid collection methods can be utilizedto collect the biological sample from the subject in order to determinethe level of DNA, RNA and/or polypeptide of the marker of interest inthe subject.

Examples include, but are not limited to, fine needle biopsy, needlebiopsy, core needle biopsy and surgical biopsy (e.g., brain biopsy), andlavage. Regardless of the procedure employed, once a biopsy/sample isobtained the level of the marker can be determined and a diagnosis canthus be made.

Determining the level of the same marker in normal tissues of the sameorigin is preferably effected along-side to detect an elevatedexpression and/or amplification and/or a decreased expression, of themarker as opposed to the normal tissues.

In some embodiments, the term “test amount” of a marker refers to anamount of a marker in a subject's sample that is consistent with adiagnosis of a particular disease or condition. A test amount can beeither in absolute amount (e.g., microgram/ml) or a relative amount(e.g., relative intensity of signals).

In some embodiments, the term “control amount” of a marker can be anyamount or a range of amounts to be compared against a test amount of amarker. For example, a control amount of a marker can be the amount of amarker in a patient with a particular disease or condition or a personwithout such a disease or condition. A control amount can be either inabsolute amount (e.g., microgram/ml) or a relative amount (e.g.,relative intensity of signals).

In some embodiments, the term “detect” refers to identifying thepresence, absence or amount of the object to be detected.

In some embodiments, the term “label” includes any moiety or itemdetectable by spectroscopic, photo chemical, biochemical,immunochemical, or chemical means. For example, useful labels include32P, 35S, fluorescent dyes, electron-dense reagents, enzymes (e.g., ascommonly used in an ELISA), biotin-streptavadin, dioxigenin, haptens andproteins for which antisera or monoclonal antibodies are available, ornucleic acid molecules with a sequence complementary to a target. Thelabel often generates a measurable signal, such as a radioactive,chromogenic, or fluorescent signal, that can be used to quantify theamount of bound label in a sample. The label can be incorporated in orattached to a primer or probe either covalently, or through ionic, vander Waals or hydrogen bonds, e.g., incorporation of radioactivenucleotides, or biotinylated nucleotides that are recognized bystreptavadin. The label may be directly or indirectly detectable.Indirect detection can involve the binding of a second label to thefirst label, directly or indirectly. For example, the label can be theligand of a binding partner, such as biotin, which is a binding partnerfor streptavadin, or a nucleotide sequence, which is the binding partnerfor a complementary sequence, to which it can specifically hybridize.The binding partner may itself be directly detectable, for example, anantibody may be itself labeled with a fluorescent molecule. The bindingpartner also may be indirectly detectable, for example, a nucleic acidhaving a complementary nucleotide sequence can be a part of a branchedDNA molecule that is in turn detectable through hybridization with otherlabeled nucleic acid molecules (see, e.g., P. D. Fahrlander and A.Klausner, Bio/Technology 6:1165 (1988)). Quantitation of the signal isachieved by, e.g., scintillation counting, densitometry, or flowcytometry.

Exemplary detectable labels, optionally and preferably for use withimmunoassays, include but are not limited to magnetic beads, fluorescentdyes, radiolabels, enzymes (e.g., horse radish peroxide, alkalinephosphatase and others commonly used in an ELISA), and calorimetriclabels such as colloidal gold or colored glass or plastic beads.Alternatively, the marker in the sample can be detected using anindirect assay, wherein, for example, a second, labeled antibody is usedto detect bound marker-specific antibody, and/or in a competition orinhibition assay wherein, for example, a monoclonal antibody which bindsto a distinct epitope of the marker are incubated simultaneously withthe mixture.

“Immunoassay” is an assay that uses an antibody to specifically bind anantigen. The immunoassay is characterized by the use of specific bindingproperties of a particular antibody to isolate, target, and/or quantifythe antigen.

The phrase “specifically (or selectively) binds” to an antibody or“specifically (or selectively) immunoreactive with,” or “specificallyinteracts or binds” when referring to a protein or peptide (or otherepitope), refers, in some embodiments, to a binding reaction that isdeterminative of the presence of the protein in a heterogeneouspopulation of proteins and other biologics. Thus, under designatedimmunoassay conditions, the specified antibodies bind to a particularprotein at least two times greater than the background (non-specificsignal) and do not substantially bind in a significant amount to otherproteins present in the sample. Specific binding to an antibody undersuch conditions may require an antibody that is selected for itsspecificity for a particular protein. For example, polyclonal antibodiesraised to seminal basic protein from specific species such as rat,mouse, or human can be selected to obtain only those polyclonalantibodies that are specifically immunoreactive with seminal basicprotein and not with other proteins, except for polymorphic variants andalleles of seminal basic protein. This selection may be achieved bysubtracting out antibodies that cross-react with seminal basic proteinmolecules from other species. A variety of immunoassay formats may beused to select antibodies specifically immunoreactive with a particularprotein. For example, solid-phase ELISA immunoassays are routinely usedto select antibodies specifically immunoreactive with a protein (see,e.g., Harlow & Lane, Antibodies, A Laboratory Manual (1988), for adescription of immunoassay formats and conditions that can be used todetermine specific immunoreactivity). Typically a specific or selectivereaction will be at least twice background signal or noise and moretypically more than 10 to 100 times background.

In another embodiment, this invention provides a method for detectingthe polypeptides of this invention in a biological sample, comprising:contacting a biological sample with an antibody specifically recognizinga polypeptide according to the present invention and detecting saidinteraction; wherein the presence of an interaction correlates with thepresence of a polypeptide in the biological sample.

In some embodiments of the present invention, the polypeptides describedherein are non-limiting examples of markers for diagnosing a diseaseand/or an indicative condition. Each marker of the present invention canbe used alone or in combination, for various uses, including but notlimited to, prognosis, prediction, screening, early diagnosis,determination of progression, therapy selection and treatment monitoringof a disease and/or an indicative condition.

In a related object the detected diseases will include cancers such asnon-solid and solid tumors, sarcomas, hematological malignanciesincluding but not limited to acute lymphocytic leukemia, chroniclymphocytic leukemia, acute myelogenous leukemia, chronic myelogenousleukemia, multiple myeloma, Hodgkin's lymphoma, Non-Hodgkin's lymphoma,cancer of the breast, prostate, lung, ovary, colon, spleen, kidney,bladder, head and neck, uterus, testicles, stomach, cervix, liver, bone,skin, pancreas, brain and wherein the cancer may be non-metastatic,invasive or metastatic.

In another related object the detected diseases will include autoimmuneand neoplastic disorders selected from the group consisting of Multiplesclerosis; Psoriasis; Rheumatoid arthritis; Systemic lupuserythematosus; Ulcerative colitis; Crohn's disease; immune disordersassociated with graft transplantation rejection, benign lymphocyticangiitis, lupus erythematosus, Hashimoto's thyroiditis, primarymyxedema, Graves' disease, pernicious anemia, autoimmune atrophicgastritis, Addison's disease, insulin dependent diabetes mellitis, goodpasture's syndrome, myasthenia gravis, pemphigus, sympatheticophthalmia, autoimmune uveitis, autoimmune hemolytic anemia, idiopathicthrombocytopenia, primary biliary cirrhosis, chronic action hepatitis,ulceratis colitis, Sjogren's syndrome, rheumatic disease, polymyositis,scleroderma, mixed connective tissue disease, inflammatory rheumatism,degenerative rheumatism, extra-articular rheumatism, collagen diseases,chronic polyarthritis, psoriasis arthropathica, ankylosing spondylitis,juvenile rheumatoid arthritis, periarthritis humeroscapularis,panarteriitis nodosa, progressive systemic scleroderma, arthritisuratica, dermatomyositis, muscular rheumatism, myositis, myogelosis andchondrocalcinosis.

In another related object the detected diseases will include rejectionof any organ transplant and/or Graft versus host disease.

Each polypeptide/polynucleotide of the present invention can be usedalone or in combination, for various uses, including but not limited to,prognosis, prediction, screening, early diagnosis, determination ofprogression, therapy selection and treatment monitoring of diseaseand/or an indicative condition, as detailed above.

Such a combination may optionally comprise any subcombination ofmarkers, and/or a combination featuring at least one other marker, forexample a known marker. Furthermore, such a combination may optionallyand preferably be used as described above with regard to determining aratio between a quantitative or semi-quantitative measurement of anymarker described herein to any other marker described herein, and/or anyother known marker, and/or any other marker.

According to further embodiments of the present invention markers of thepresent invention might optionally be used alone or in combination withknown markers for lung cancer, including but not limited to CEA, CA15-3,Beta-2-microglobulin, CA19-9, TPA, and/or in combination with the knownproteins for the variant marker as described herein.

According to further embodiments of the present invention markers of thepresent invention might optionally be used alone or in combination withknown markers for ovarian cancer, including but not limited to CEA,CA125 (Mucin 16), CA72-4TAG, CA-50, CA 54-61, CA-195 and CA 19-9 incombination with CA-125, and/or in combination with the known proteinsfor the variant marker as described herein.

According to further embodiments of the present invention markers of thepresent invention might optionally be used alone or in combination withknown markers for colon cancer, including but not limited to CEA,CA19-9, CA50, and/or in combination with the known proteins for thevariant marker as described herein.

In some embodiments of the present invention, there are provided ofmethods, uses, devices and assays for the diagnosis of a disease orcondition. Optionally a plurality of markers may be used with thepresent invention. The plurality of markers may optionally include amarkers described herein, and/or one or more known markers. Theplurality of markers is preferably then correlated with the disease orcondition. For example, such correlating may optionally comprisedetermining the concentration of each of the plurality of markers, andindividually comparing each marker concentration to a threshold level.Optionally, if the marker concentration is above or below the thresholdlevel (depending upon the marker and/or the diagnostic test beingperformed), the marker concentration correlates with the disease orcondition. Optionally and preferably, a plurality of markerconcentrations correlates with the disease or condition.

Alternatively, such correlating may optionally comprise determining theconcentration of each of the plurality of markers, calculating a singleindex value based on the concentration of each of the plurality ofmarkers, and comparing the index value to a threshold level.

Also alternatively, such correlating may optionally comprise determininga temporal change in at least one of the markers, and wherein thetemporal change is used in the correlating step.

Also alternatively, such correlating may optionally comprise determiningwhether at least “X” number of the plurality of markers has aconcentration outside of a predetermined range and/or above or below athreshold (as described above). The value of “X” may optionally be onemarker, a plurality of markers or all of the markers; alternatively oradditionally, rather than including any marker in the count for “X”, oneor more specific markers of the plurality of markers may optionally berequired to correlate with the disease or condition (according to arange and/or threshold).

Also alternatively, such correlating may optionally comprise determiningwhether a ratio of marker concentrations for two markers is outside arange and/or above or below a threshold. Optionally, if the ratio isabove or below the threshold level and/or outside a range, the ratiocorrelates with the disease or condition.

Optionally, a combination of two or more these correlations may be usedwith a single panel and/or for correlating between a plurality ofpanels.

Optionally, the method distinguishes a disease or condition with asensitivity of at least 70% at a specificity of at least 85% whencompared to normal subjects. As used herein, sensitivity relates to thenumber of positive (diseased) samples detected out of the total numberof positive samples present; specificity relates to the number of truenegative (non-diseased) samples detected out of the total number ofnegative samples present. Preferably, the method distinguishes a diseaseor condition with a sensitivity of at least 80% at a specificity of atleast 90% when compared to normal subjects. More preferably, the methoddistinguishes a disease or condition with a sensitivity of at least 90%at a specificity of at least 90% when compared to normal subjects. Alsomore preferably, the method distinguishes a disease or condition with asensitivity of at least 70% at a specificity of at least 85% whencompared to subjects exhibiting symptoms that mimic disease or conditionsymptoms.

A marker panel may be analyzed in a number of fashions well known tothose of skill in the art. For example, each member of a panel may becompared to a “normal” value, or a value indicating a particularoutcome. A particular diagnosis/prognosis may depend upon the comparisonof each marker to this value; alternatively, if only a subset of markersis outside of a normal range, this subset may be indicative of aparticular diagnosis/prognosis. The skilled artisan will also understandthat diagnostic markers, differential diagnostic markers, prognosticmarkers, time of onset markers, disease or condition differentiatingmarkers, etc., may be combined in a single assay or device. Markers mayalso be commonly used for multiple purposes by, for example, applying adifferent threshold or a different weighting factor to the marker forthe different purposes.

In one embodiment, the panels comprise markers for the followingpurposes: diagnosis of a disease; diagnosis of disease and indication ifthe disease is in an acute phase and/or if an acute attack of thedisease has occurred; diagnosis of disease and indication if the diseaseis in a non-acute phase and/or if a non-acute attack of the disease hasoccurred; indication whether a combination of acute and non-acute phasesor attacks has occurred; diagnosis of a disease and prognosis of asubsequent adverse outcome; diagnosis of a disease and prognosis of asubsequent acute or non-acute phase or attack; disease progression (forexample for cancer, such progression may include for example occurrenceor recurrence of metastasis).

The above diagnoses may also optionally include differential diagnosisof the disease to distinguish it from other diseases, including thosediseases that may feature one or more similar or identical symptoms.

In certain embodiments, one or more diagnostic or prognostic indicatorsare correlated to a condition or disease by merely the presence orabsence of the indicators. In other embodiments, threshold levels of adiagnostic or prognostic indicators can be established, and the level ofthe indicators in a patient sample can simply be compared to thethreshold levels. The sensitivity and specificity of a diagnostic and/orprognostic test depends on more than just the analytical “quality” ofthe test—they also depend on the definition of what constitutes anabnormal result. In practice, Receiver Operating Characteristic curves,or “ROC” curves, are typically calculated by plotting the value of avariable versus its relative frequency in “normal” and “disease”populations, and/or by comparison of results from a subject before,during and/or after treatment.

According to embodiments of the present invention, VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, or FXYD3 protein, polynucleotide or afragment thereof, may be featured as a biomarker for detecting diseaseand/or an indicative condition, as detailed above.

According to still other embodiments, the present invention optionallyand preferably encompasses any amino acid sequence or fragment thereofencoded by a nucleic acid sequence corresponding to VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, or FXYD3 as described herein. Anyoligopeptide or peptide relating to such an amino acid sequence orfragment thereof may optionally also (additionally or alternatively) beused as a biomarker.

In still other embodiments, the present invention provides a method fordetecting a polynucleotide of this invention in a biological sample,using NAT based assays, comprising: hybridizing the isolated nucleicacid molecules or oligonucleotide fragments of at least about a minimumlength to a nucleic acid material of a biological sample and detecting ahybridization complex; wherein the presence of a hybridization complexcorrelates with the presence of the polynucleotide in the biologicalsample. Non-limiting examples of methods or assays are described below.

The present invention also relates to kits based upon such diagnosticmethods or assays.

Nucleic Acid Technology (NAT) Based Assays:

Detection of a nucleic acid of interest in a biological sample may alsooptionally be effected by NAT-based assays, which involve nucleic acidamplification technology, such as PCR for example (or variations thereofsuch as real-time PCR for example). As used herein, a “primer” definesan oligonucleotide which is capable of annealing to (hybridizing with) atarget sequence, thereby creating a double stranded region which canserve as an initiation point for DNA synthesis under suitableconditions. Amplification of a selected, or target, nucleic acidsequence may be carried out by a number of suitable methods known in theart. Non-limiting examples of amplification techniques includepolymerase chain reaction (PCR), ligase chain reaction (LCR), stranddisplacement amplification (SDA), transcription-based amplification, theq3 replicase system and NASBA (Kwoh et al., 1989, Proc. Natl. Acad. Sci.USA 86, 1173-1177; Lizardi et al., 1988, BioTechnology 6:1197-1202;Malek et al., 1994, Methods Mol. Biol., 28:253-260; and Sambrook et al.,1989, supra). Non-limiting examples of Nucleic Acid Technology-basedassay is selected from the group consisting of a PCR, Real-Time PCR,LCR, Self-Sustained Synthetic Reaction, Q-Beta Replicase, Cycling probereaction, Branched DNA, RFLP analysis, DGGE/TGGE, Single-StrandConformation Polymorphism, Dideoxy fingerprinting, microarrays,Fluorescense In Situ Hybridization and Comparative GenomicHybridization. The terminology “amplification pair” (or “primer pair”)refers herein to a pair of oligonucleotides (oligos) of the presentinvention, which are selected to be used together in amplifying aselected nucleic acid sequence by one of a number of types ofamplification processes, preferably a polymerase chain reaction. Ascommonly known in the art, the oligos are designed to bind to acomplementary sequence under selected conditions. In one particularembodiment, amplification of a nucleic acid sample from a patient isamplified under conditions which favor the amplification of the mostabundant differentially expressed nucleic acid. In one preferredembodiment, RT-PCR is carried out on an mRNA sample from a patient underconditions which favor the amplification of the most abundant mRNA. Inanother preferred embodiment, the amplification of the differentiallyexpressed nucleic acids is carried out simultaneously. It will berealized by a person skilled in the art that such methods could beadapted for the detection of differentially expressed proteins insteadof differentially expressed nucleic acid sequences. The nucleic acid(i.e. DNA or RNA) for practicing the present invention may be obtainedaccording to well known methods.

Oligonucleotide primers of the present invention may be of any suitablelength, depending on the particular assay format and the particularneeds and targeted genomes employed. Optionally, the oligonucleotideprimers are at least 12 nucleotides in length, preferably between 15 and24 molecules, and they may be adapted to be especially suited to achosen nucleic acid amplification system. As commonly known in the art,the oligonucleotide primers can be designed by taking into considerationthe melting point of hybridization thereof with its targeted sequence(Sambrook et al., 1989, Molecular Cloning—A Laboratory Manual, 2ndEdition, CSH Laboratories; Ausubel et al., 1989, in Current Protocols inMolecular Biology, John Wiley & Sons Inc., N.Y.).

Immunoassays

In another embodiment of the present invention, an immunoassay can beused to qualitatively or quantitatively detect and analyze markers in asample. This method comprises: providing an antibody that specificallybinds to a marker; contacting a sample with the antibody; and detectingthe presence of a complex of the antibody bound to the marker in thesample.

To prepare an antibody that specifically binds to a marker, purifiedprotein markers can be used. Antibodies that specifically bind to aprotein marker can be prepared using any suitable methods known in theart.

After the antibody is provided, a marker can be detected and/orquantified using any of a number of well recognized immunologicalbinding assays. Useful assays include, for example, an enzyme immuneassay (EIA) such as enzyme-linked immunosorbent assay (ELISA), aradioimmune assay (RIA), a Western blot assay, or a slot blot assay see,e.g., U.S. Pat. Nos. 4,366,241; 4,376,110; 4,517,288; and 4,837,168).Generally, a sample obtained from a subject can be contacted with theantibody that specifically binds the marker.

Optionally, the antibody can be fixed to a solid support to facilitatewashing and subsequent isolation of the complex, prior to contacting theantibody with a sample. Examples of solid supports include but are notlimited to glass or plastic in the form of, e.g., a microtiter plate, astick, a bead, or a microbead. Antibodies can also be attached to asolid support.

After incubating the sample with antibodies, the mixture is washed andthe antibody-marker complex formed can be detected. This can beaccomplished by incubating the washed mixture with a detection reagent.Alternatively, the marker in the sample can be detected using anindirect assay, wherein, for example, a second, labeled antibody is usedto detect bound marker-specific antibody, and/or in a competition orinhibition assay wherein, for example, a monoclonal antibody which bindsto a distinct epitope of the marker are incubated simultaneously withthe mixture.

Throughout the assays, incubation and/or washing steps may be requiredafter each combination of reagents. Incubation steps can vary from about5 seconds to several hours, preferably from about 5 minutes to about 24hours. However, the incubation time will depend upon the assay format,marker, volume of solution, concentrations and the like. Usually theassays will be carried out at ambient temperature, although they can beconducted over a range of temperatures, such as 10° C. to 40° C.

The immunoassay can be used to determine a test amount of a marker in asample from a subject. First, a test amount of a marker in a sample canbe detected using the immunoassay methods described above. If a markeris present in the sample, it will form an antibody-marker complex withan antibody that specifically binds the marker under suitable incubationconditions described above. The amount of an antibody-marker complex canoptionally be determined by comparing to a standard. As noted above, thetest amount of marker need not be measured in absolute units, as long asthe unit of measurement can be compared to a control amount and/orsignal.

Radio-immunoassay (RIA): In one version, this method involvesprecipitation of the desired substrate and in the methods detailedherein below, with a specific antibody and radiolabeled antibody bindingprotein (e.g., protein A labeled with I125) immobilized on aprecipitable carrier such as agarose beads. The number of counts in theprecipitated pellet is proportional to the amount of substrate.

In an alternate version of the RIA, a labeled substrate and anunlabelled antibody binding protein are employed. A sample containing anunknown amount of substrate is added in varying amounts. The decrease inprecipitated counts from the labeled substrate is proportional to theamount of substrate in the added sample.

Enzyme linked immunosorbent assay (ELISA): This method involves fixationof a sample (e.g., fixed cells or a proteinaceous solution) containing aprotein substrate to a surface such as a well of a microtiter plate. Asubstrate specific antibody coupled to an enzyme is applied and allowedto bind to the substrate. Presence of the antibody is then detected andquantitated by a colorimetric reaction employing the enzyme coupled tothe antibody. Enzymes commonly employed in this method includehorseradish peroxidase and alkaline phosphatase. If well calibrated andwithin the linear range of response, the amount of substrate present inthe sample is proportional to the amount of color produced. A substratestandard is generally employed to improve quantitative accuracy.

Western blot: This method involves separation of a substrate from otherprotein by means of an acrylamide gel followed by transfer of thesubstrate to a membrane (e.g., nylon or PVDF). Presence of the substrateis then detected by antibodies specific to the substrate, which are inturn detected by antibody binding reagents. Antibody binding reagentsmay be, for example, protein A, or other antibodies. Antibody bindingreagents may be radiolabeled or enzyme linked as described hereinabove.Detection may be by autoradiography, colorimetric reaction orchemiluminescence. This method allows both quantitation of an amount ofsubstrate and determination of its identity by a relative position onthe membrane which is indicative of a migration distance in theacrylamide gel during electrophoresis.

Immunohistochemical analysis: This method involves detection of asubstrate in situ in fixed cells by substrate specific antibodies. Thesubstrate specific antibodies may be enzyme linked or linked tofluorophores. Detection is by microscopy and subjective evaluation. Ifenzyme linked antibodies are employed, a colorimetric reaction may berequired.

Fluorescence activated cell sorting (FACS): This method involvesdetection of a substrate in situ in cells by substrate specificantibodies. The substrate specific antibodies are linked tofluorophores. Detection is by means of a cell sorting machine whichreads the wavelength of light emitted from each cell as it passesthrough a light beam. This method may employ two or more antibodiessimultaneously.

Radio-Imaging Methods

These methods include but are not limited to, positron emissiontomography (PET) single photon emission computed tomography (SPECT).Both of these techniques are non-invasive, and can be used to detectand/or measure a wide variety of tissue events and/or functions, such asdetecting cancerous cells for example. Unlike PET, SPECT can optionallybe used with two labels simultaneously. SPECT has some other advantagesas well, for example with regard to cost and the types of labels thatcan be used. For example, U.S. Pat. No. 6,696,686 describes the use ofSPECT for detection of breast cancer, and is hereby incorporated byreference as if fully set forth herein.

Theranostics:

The term theranostics describes the use of diagnostic testing todiagnose the disease, choose the correct treatment regime according tothe results of diagnostic testing and/or monitor the patient response totherapy according to the results of diagnostic testing. Theranostictests can be used to select patients for treatments that areparticularly likely to benefit them and unlikely to produceside-effects. They can also provide an early and objective indication oftreatment efficacy in individual patients, so that (if necessary) thetreatment can be altered with a minimum of delay. For example: DAKO andGenentech together created HercepTest and Herceptin (trastuzumab) forthe treatment of breast cancer, the first theranostic test approvedsimultaneously with a new therapeutic drug. In addition to HercepTest(which is an immunohistochemical test), other theranostic tests are indevelopment which use traditional clinical chemistry, immunoassay,cell-based technologies and nucleic acid tests. PPGx's recently launchedTPMT (thiopurine S-methyltransferase) test, which is enabling doctors toidentify patients at risk for potentially fatal adverse reactions to6-mercaptopurine, an agent used in the treatment of leukemia. Also, NovaMolecular pioneered SNP genotyping of the apolipoprotein E gene topredict Alzheimer's disease patients' responses to cholinomimetictherapies and it is now widely used in clinical trials of new drugs forthis indication. Thus, the field of theranostics represents theintersection of diagnostic testing information that predicts theresponse of a patient to a treatment with the selection of theappropriate treatment for that particular patient.

Surrogate Markers:

A surrogate marker is a marker, that is detectable in a laboratoryand/or according to a physical sign or symptom on the patient, and thatis used in therapeutic trials as a substitute for a clinicallymeaningful endpoint. The surrogate marker is a direct measure of how apatient feels, functions, or survives which is expected to predict theeffect of the therapy. The need for surrogate markers mainly arises whensuch markers can be measured earlier, more conveniently, or morefrequently than the endpoints of interest in terms of the effect of atreatment on a patient, which are referred to as the clinical endpoints.Ideally, a surrogate marker should be biologically plausible, predictiveof disease progression and measurable by standardized assays (includingbut not limited to traditional clinical chemistry, immunoassay,cell-based technologies, nucleic acid tests and imaging modalities).

Surrogate endpoints were used first mainly in the cardiovascular area.For example, antihypertensive drugs have been approved based on theireffectiveness in lowering blood pressure. Similarly, in the past,cholesterol-lowering agents have been approved based on their ability todecrease serum cholesterol, not on the direct evidence that theydecrease mortality from atherosclerotic heart disease. The measurementof cholesterol levels is now an accepted surrogate marker ofatherosclerosis. In addition, currently two commonly used surrogatemarkers in HIV studies are CD4+ T cell counts and quantitative plasmaHIV RNA (viral load). In some embodiments of this invention, thepolypeptide/polynucleotide expression pattern may serve as a surrogatemarker for a particular disease, as will be appreciated by one skilledin the art.

Uses and Methods of the Invention

The VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3 drugs accordingto the invention, especially antibodies, particularly the humanantibodies, antibody compositions, and soluble conjugates containing theectodomain of the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3antigen or a fragment or variant thereof, or a corresponding nucleicacid sequence or vector or cell expressing same and methods of thepresent invention have numerous in vitro and in vivo diagnostic andtherapeutic utilities involving the diagnosis and treatment of VSIG1,ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3 antigen related disordersand/or disorders wherein modulation of immune co-stimulation e.g.,involving B7-related immune costimulation involving VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 antigen is therapeuticallydesirable. As noted these conditions include in particular cancers thatdifferentially express the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32,or FXYD3 antigen such as lung cancer, ovarian cancer, colon cancer,including invasive and metastatic forms thereof, and/or autoimmuneconditions wherein modulation of costimulation such as involving B7 istherapeutically desirable. The subject anti-VSIG1, anti-ILDR1,anti-LOC253012, anti-AI216611, anti-C1ORF32 or anti-FXYD3 antibodies mayprevent B7 mediated negative stimulation of T cell activity againstcancer cells and/or prevent positive stimulation of T cell activity.Such antibodies may be used in the treatment of conditions includingcancers such non-solid and solid tumors, sarcomas, hematologicalmalignancies including but not limited to acute lymphocytic leukemia,chronic lymphocytic leukemia, acute myelogenous leukemia, chronicmyelogenous leukemia, multiple myeloma, Hodgkin's lymphoma,Non-Hodgkin's lymphoma, cancer of the breast, prostate, lung, ovary,colon, spleen, kidney, bladder, head and neck, uterus, testicles,stomach, cervix, liver, bone, skin, pancreas, brain and wherein thecancer may be non-metastatic, invasive or metastatic as well asnon-malignant disorders such as immune disorders including but notlimited to transplant rejection and graft versus host disease, andautoimmune disorders such as afore-mentioned.

For example, these molecules can be administered to cells in culture, invitro or ex vivo, or to human subjects, e.g., in vivo, to treat, preventand to diagnose a variety of disorders. Preferred subjects include humanpatients having disorders mediated by cells expressing the VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, or FXYD3 antigen and cells that possesVSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 activity. Themethods are particularly suitable for treating human patients having adisorder associated with aberrant VSIG1, ILDR1, LOC253012, AI216611,C1ORF32 or FXYD3 antigen expression using antibodies that specificallybind AI581519_P3 (SEQ ID NO:11), AI581519_P4 (SEQ ID NO:12), AI581519_P5(SEQ ID NO:13), AI581519_P7 (SEQ ID NO:14), AI581519_P9 (SEQ ID NO:15),AI581519_P10 (SEQ ID NO:16), AA424839_P3 (SEQ ID NO:22), AA424839_P5(SEQ ID NO:21), AA424839_P7 (SEQ ID NO:23), or AA424839_(—)1_P11 (SEQ IDNO:24), H68654_(—)1_P2 (SEQ ID NO:35), H68654_(—)1_P5 (SEQ ID NO:36),H68654_(—)1_P7 (SEQ ID NO:37), H68654_(—)1_P12 (SEQ ID NO:38),H68654_(—)1_P13 (SEQ ID NO:39), H68654_(—)1_P14 (SEQ ID NO:40),AI216611_P0 (SEQ ID NO:43), AI216611_P1 (SEQ ID NO:44), H19011_(—)1_P8(SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), R31375_P0 (SEQ ID NO:70),R31375_P14 (SEQ ID NO:72), R31375_P31 (SEQ ID NO:73) or R31375_P33 (SEQID NO:74).

VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, or FXYD3 drugs according tothe invention, are administered together with another agent, the two canbe administered in either order or simultaneously.

Given the specific binding of the antibodies of the invention for VSIG1,ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 the antibodies of theinvention can be used to specifically detect VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 or FXYD3 expression on the surface of cells and,moreover, can be used to purify VSIG1, ILDR1, LOC253012, AI216611,C1ORF32 or FXYD3 antigen via immunoaffinity purification.

Furthermore, given the expression of VSIG1, ILDR1, LOC253012, AI216611,C1ORF32 or FXYD3 on various tumor cells, the human antibodies, antibodycompositions and methods of the present invention can be used to treat asubject with a tumorigenic disorder, e.g., a disorder characterized bythe presence of tumor cells expressing VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 or FXYD3 antigen such as lung cancer and ovariancancer, as mentioned.

In one embodiment, the antibodies (e.g., human monoclonal antibodies,multispecific and bispecific molecules and compositions) of theinvention can be used to detect levels of VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 or FXYD3 or levels of cells which contain VSIG1,ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3, respectively, on theirmembrane surface, which levels can then be linked to certain diseasesymptoms. Alternatively, the antibodies can be used to inhibit or blockVSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 function which, inturn, can be linked to the prevention or amelioration of certain diseasesymptoms, thereby implicating VSIG1, ILDR1, LOC253012, AI216611, C1ORF32or FXYD3, respectively, as a mediator of the disease. This can beachieved by contacting a sample and a control sample with theanti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32 oranti-FXYD3 antibody under conditions that allow for the formation of acomplex between the corresponding antibody and VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 or FXYD3, respectively. Any complexes formed betweenthe antibody and VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 aredetected and compared in the sample and the control.

In another embodiment, the antibodies (e.g., human antibodies,multispecific and bispecific molecules and compositions) of theinvention can be initially tested for binding activity associated withtherapeutic or diagnostic use in vitro. For example, compositions of theinvention can be tested using low cytometric assays.

The antibodies (e.g., human antibodies, multispecific and bispecificmolecules, immunoconjugates and compositions) of the invention haveadditional utility in therapy and diagnosis of VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 or FXYD3-related diseases. For example, the humanmonoclonal antibodies, the multispecific or bispecific molecules and theimmunoconjugates can be used to elicit in vivo or in vitro one or moreof the following biological activities: to inhibit the growth of and/orkill a cell expressing VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 orFXYD3; to mediate phagocytosis or ADCC of a cell expressing VSIG1,ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 in the presence of humaneffector cells, or to block VSIG1, ILDR1, LOC253012, AI216611, C1ORF32or FXYD3 ligand binding to VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 orFXYD3, respectively.

In a particular embodiment, the antibodies (e.g., human antibodies,multispecific and bispecific molecules and compositions) are used invivo to treat, prevent or diagnose a variety of VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 or FXYD3-related diseases. Examples of VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3-related diseases include, amongothers, cancer, such as lung cancer, ovarian cancer, colon cancer, othernon-solid and solid tumors, sarcomas, hematological malignanciesincluding but not limited to acute lymphocytic leukemia, chroniclymphocytic leukemia, acute myelogenous leukemia, chronic myelogenousleukemia, multiple myeloma, Hodgkin's lymphoma, Non-Hodgkin's lymphoma,cancer of the breast, prostate, spleen, kidney, bladder, head and neck,uterus, testicles, stomach, cervix, liver, bone, skin, pancreas, brainand wherein the cancer may be non-metastatic, invasive or metastatic.Additional examples of VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 orFXYD3-related diseases include, among others, non-malignant disorderssuch as immune disorders including but not limited to autoimmunediseases, transplant rejection and graft versus host disease. Suchdisorders include by way of example autoimmune diseases selected frommultiple sclerosis; psoriasis; rheumatoid arthritis; Systemic lupuserythematosus; Ulcerative colitis; Crohn's disease, immune disordersassociated with graft transplantation rejection, benign lymphocyticangiitis, lupus erythematosus, Hashimoto's thyroiditis, primarymyxedema, Graves' disease, pernicious anemia, autoimmune atrophicgastritis, Addison's disease, insulin dependent diabetes mellitis, Goodpasture's syndrome, myasthenia gravis, pemphigus, sympatheticophthalmia, autoimmune uveitis, autoimmune hemolytic anemia, idiopathicthrombocytopenia, primary biliary cirrhosis, chronic action hepatitis,ulceratis colitis, Sjogren's syndrome, rheumatic disease, polymyositis,scleroderma, mixed connective tissue disease, inflammatory rheumatism,degenerative rheumatism, extra-articular rheumatism, collagen diseases,chronic polyarthritis, psoriasis arthropathica, ankylosing spondylitis,juvenile rheumatoid arthritis, periarthritis humeroscapularis,panarteriitis nodosa, progressive systemic scleroderma, arthritisuratica, dermatomyositis, muscular rheumatism, myositis, myogelosis andchondrocalcinosis.

Suitable routes of administering the antibody compositions (e.g., humanmonoclonal antibodies, multispecific and bispecific molecules andimmunoconjugates) of the invention in vivo and in vitro are well knownin the art and can be selected by those of ordinary skill. For example,the antibody compositions can be administered by injection (e.g.,intravenous or subcutaneous). Suitable dosages of the molecules usedwill depend on the age and weight of the subject and the concentrationand/or formulation of the antibody composition.

As previously described, human anti-VSIG1, anti-ILDR1, anti-LOC253012,anti-AI216611, anti-C1ORF32 or anti-FXYD3 antibodies of the inventioncan be co-administered with one or other more therapeutic agents, e.g.,an cytotoxic agent, a radiotoxic agent or an immunosuppressive agent.The antibody can be linked to the agent (as an immunocomplex) or can beadministered separate from the agent. In the latter case (separateadministration), the antibody can be administered before, after orconcurrently with the agent or can be co-administered with other knowntherapies, e.g., an anti-cancer therapy, e.g., radiation. Suchtherapeutic agents include, among others, anti-neoplastic agents such asdoxorubicin (adriamycin), cisplatin bleomycin sulfate, carmustine,chlorambucil, and cyclophosphamide hydroxyurea which, by themselves, areonly effective at levels which are toxic or subtoxic to a patient.Cisplatin is intravenously administered as a 100 mg/dose once every fourweeks and adriamycin is intravenously administered as a 60-75 mg/ml doseonce every 21 days. Co-administration of the human anti-VSIG1,anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32 or anti-FXYD3antibodies, or antigen binding fragments thereof, of the presentinvention with chemotherapeutic agents provides two anti-cancer agentswhich operate via different mechanisms which yield a cytotoxic effect tohuman tumor cells. Such co-administration can solve problems due todevelopment of resistance to drugs or a change in the antigenicity ofthe tumor cells which would render them unreactive with the antibody.

Target-specific effector cells, e.g., effector cells linked tocompositions (e.g., human antibodies, multispecific and bispecificmolecules) of the invention can also be used as therapeutic agents.Effector cells for targeting can be human leukocytes such asmacrophages, neutrophils or monocytes. Other cells include eosinophils,natural killer cells and other IgG- or IgA-receptor bearing cells. Ifdesired, effector cells can be obtained from the subject to be treated.The target-specific effector cells can be administered as a suspensionof cells in a physiologically acceptable solution. The number of cellsadministered can be in the order of 10−8 to 10−9 but will vary dependingon the therapeutic purpose. In general, the amount will be sufficient toobtain localization at the target cell, e.g., a tumor cell expressingVSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 and to effect cellkilling by, e.g., phagocytosis. Routes of administration can also vary.

Therapy with target-specific effector cells can be performed inconjunction with other techniques for removal of targeted cells. Forexample, anti-tumor therapy using the compositions (e.g., humanantibodies, multispecific and bispecific molecules) of the inventionand/or effector cells armed with these compositions can be used inconjunction with chemotherapy. Additionally, combination immunotherapymay be used to direct two distinct cytotoxic effector populations towardtumor cell rejection. For example, anti-VSIG1, anti-ILDR1,anti-LOC253012, anti-AI216611, anti-C1ORF32 or anti-FXYD3 antibodieslinked to anti-Fc-gamma R1 or anti-CD3 may be used in conjunction withIgG- or IgA-receptor specific binding agents.

Bispecific and multispecific molecules of the invention can also be usedto modulate FcgammaR or FcgammaR levels on effector cells, such as bycapping and elimination of receptors on the cell surface. Mixtures ofanti-Fc receptors can also be used for this purpose.

The compositions (e.g., human antibodies, multispecific and bispecificmolecules and immunoconjugates) of the invention which have complementbinding sites, such as portions from IgG1, -2, or -3 or IgM which bindcomplement, can also be used in the presence of complement. In oneembodiment, ex vivo treatment of a population of cells comprising targetcells with a binding agent of the invention and appropriate effectorcells can be supplemented by the addition of complement or serumcontaining complement. Phagocytosis of target cells coated with abinding agent of the invention can be improved by binding of complementproteins. In another embodiment target cells coated with thecompositions (e.g., human antibodies, multispecific and bispecificmolecules) of the invention can also be lysed by complement. In yetanother embodiment, the compositions of the invention do not activatecomplement.

The compositions (e.g., human antibodies, multispecific and bispecificmolecules and immunoconjugates) of the invention can also beadministered together with complement. Accordingly, within the scope ofthe invention are compositions comprising human antibodies,multispecific or bispecific molecules and serum or complement. Thesecompositions are advantageous in that the complement is located in closeproximity to the human antibodies, multispecific or bispecificmolecules. Alternatively, the human antibodies, multispecific orbispecific molecules of the invention and the complement or serum can beadministered separately.

Also within the scope of the present invention are kits comprising theVSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 antigen or VSIG1,ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 conjugates or antibodycompositions of the invention (e.g., human antibodies, bispecific ormultispecific molecules, or immunoconjugates) and instructions for use.The kit can further contain one or more additional reagents, such as animmunosuppressive reagent, a cytotoxic agent or a radiotoxic agent, orone or more additional human antibodies of the invention (e.g., a humanantibody having a complementary activity which binds to an epitope inthe VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 antigen distinctfrom the first human antibody).

Accordingly, patients treated with antibody compositions of theinvention can be additionally administered (prior to, simultaneouslywith, or following administration of a human antibody of the invention)with another therapeutic agent, such as a cytotoxic or radiotoxic agent,which enhances or augments the therapeutic effect of the humanantibodies.

In other embodiments, the subject can be additionally treated with anagent that modulates, e.g., enhances or inhibits, the expression oractivity of Fcy or Fcy receptors by, for example, treating the subjectwith a cytokine. Preferred cytokines for administration during treatmentwith the multispecific molecule include of granulocytecolony-stimulating factor (G-CSF), granulocyte-macrophagecolony-stimulating factor (GM-CSF), interferon-.gamma. (IFN-.gamma.),and tumor necrosis factor (TNF).

The compositions (e.g., human antibodies, multispecific and bispecificmolecules) of the invention can also be used to target cells expressingFc gamma R or VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3, forexample for labeling such cells. For such use, the binding agent can belinked to a molecule that can be detected. Thus, the invention providesmethods for localizing ex vivo or in vitro cells expressing Fcreceptors, such as FcgammaR, or VSIG1, ILDR1, LOC253012, AI216611,C1ORF32 or FXYD3 antigen. The detectable label can be, e.g., aradioisotope, a fluorescent compound, an enzyme, or an enzyme co-factor.

In a particular embodiment, the invention provides methods for detectingthe presence of VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3antigen in a sample, or measuring the amount of VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 or FXYD3 antigen, respectively, comprising contactingthe sample, and a control sample, with a human monoclonal antibody, oran antigen binding portion thereof, which specifically binds to VSIG1,ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3, respectively, underconditions that allow for formation of a complex between the antibody orportion thereof and VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3.The formation of a complex is then detected, wherein a differencecomplex formation between the sample compared to the control sample isindicative the presence of VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 orFXYD3 antigen in the sample. As noted the invention in particularembraces assays for detecting VSIG1, ILDR1, LOC253012, AI216611, C1ORF32or FXYD3 antigen in vitro and in vivo such as immunoassays,radioimmunoassays, radioassays, radioimaging assays, ELISAs, Westernblot, FACS, slot blot, immunohistochemical assays, and other assays wellknown to those skilled in the art.

In other embodiments, the invention provides methods for treating anVSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 mediated disorder ina subject, e.g., cancer, such as non-solid and solid tumors, sarcomas,hematological malignancies including but not limited to acutelymphocytic leukemia, chronic lymphocytic leukemia, acute myelogenousleukemia, chronic myelogenous leukemia, multiple myeloma, Hodgkin'slymphoma, Non-Hodgkin's lymphoma, cancer of the breast, prostate, lung,ovary, colon, spleen, kidney, bladder, head and neck, uterus, testicles,stomach, cervix, liver, bone, skin, pancreas, brain and wherein thecancer may be non-metastatic, invasive or metastatic, as well asnon-malignant disorders such as immune disorders including but notlimited to transplant rejection and graft versus host disease, or anautoimmune disease selected from those aforementioned and methods oftreating any condition wherein modulation of immune costimulation thatinvolves VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 istherapeutically desirable using anti-VSIG1, anti-ILDR1, anti-LOC253012,anti-AI216611, anti-C1ORF32 or anti-FXYD3 antibodies or soluble VSIG1,ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 antigen conjugates or otherdrugs that target and modulate (promote or inhibit) one or more VSIG1,ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 biological activities.

By administering the anti-VSIG1, anti-ILDR1, anti-LOC253012,anti-AI216611, anti-C1ORF32 or anti-FXYD3 antibody, soluble VSIG1,ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 antigen conjugate or otherdrug that targets the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 orFXYD3 antigen or a portion thereof to a subject, the ability of VSIG1,ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 antigen to induce suchactivities is inhibited or promoted and, thus, the associated disorderis treated. The soluble VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 orFXYD3 antigen or antigen conjugate or anti-VSIG1, anti-ILDR1,anti-LOC253012, anti-AI216611, anti-C1ORF32 or anti-FXYD3 antibody orfragment containing composition or other drug that targets and modulatesVSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3, can be administeredalone or along with another therapeutic agent, such as a cytotoxic or aradiotoxic agent which acts in conjunction with or synergistically withthe antibody composition to treat or prevent the VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 antigen mediated disease.

In yet another embodiment, immunoconjugates of the invention can be usedto target compounds (e.g., therapeutic agents, labels, cytotoxins,radiotoxins immunosuppressants, etc.) to cells which have VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 cell surface receptors by linkingsuch compounds to the antibody. Thus, the invention also providesmethods for localizing ex vivo or in vivo cells expressing VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 (e.g., with a detectable label,such as a radioisotope, a fluorescent compound, an enzyme, or an enzymeco-factor). Alternatively, the immunoconjugates can be used to killcells which have VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3cell surface receptors by targeting cytotoxins or radiotoxins to VSIG1,ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 antigen.

The present invention is further illustrated by the following sequencecharacterization of a DNA transcript encoding the VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 antigen, its domains andexpression data in normal and cancerous tissues as well as propheticexamples describing the manufacture of fully human antibodies thereto.This information and examples is illustrative and should not beconstrued as further limiting. The contents of all figures and allreferences, patents and published patent applications cited throughoutthis application are expressly incorporated herein by reference.

EXAMPLES Example 1 Methods Used to Analyze the Expression of the RNAEncoding the Proteins of the Invention

The targets of the present invention were tested with regard to theirexpression in various cancerous and non-cancerous tissue samples and/orwith regard to its expression in a wide panel of human samples whichcontains various types of immune cells, and hematological malignanciessamples and cell lines, as well as several samples of normal tissues.The list of the blood specific RNA samples used for the qRT-PCR analysisis provided in Table 1 below. A description of the samples used in thenormal tissue panel is provided in Table 2. A description of the samplesused in the lung cancer testing panel is provided in Table 3 below. Adescription of the samples used in the ovary cancer testing panel isprovided in Table 4 below. A description of the samples used in thecolon cancer testing panel is provided in Table 5 below. The keys forthe table 3, 4 and 5 are given in tables 3_(—)1, 4_(—)1, and 5_(—)1,respectively. Tests were then performed as described in the “Materialsand Experimental Procedures” section below.

TABLE 1 Samples in blood specific panel Blood panel sample DescriptionOrgan/Cell type Tumor Type 1_PBMC2 PBMCs blood-derived cells 2_PBMC3PBMCs blood-derived cells 3_Bcell1 B cells blood-derived cells 4_Bcell2B cells blood-derived cells 5_J_Bcell B cells blood-derived cells6_K_Bcells_act Bcells activated blood-derived cells 7_Tcell1 T cellsblood-derived cells 8_Tcell2 T cells blood-derived cells 9_M_CD8 CD4+ Tcells blood-derived cells 10_G_CD4_unt CD8+ T cells blood-derived cells11_H_CD4_Beads CD4+ w Activation blood-derived cells beads12_I_CD4_Beads_IL12 CD4 w act. blood-derived cells Beads + IL1213_95_CD4+CD25− CD4+CD25− blood-derived cells 15_NK NK cellsblood-derived cells 16_CD34+_1548 CD34+(PCBM1548) blood-derived cells17_CD34+_1028 CD34+(PCBM1028) blood-derived cells 18_PMN PMNsblood-derived cells 19_A_Mono Monocytes blood-derived cells20_B_Macro_imma Macrophages blood-derived cells immature 21_C_Macro_matMacrophages blood-derived cells mature 22_D_DCs_immat DCs immatureblood-derived cells 23_E_DCs_mat_LPS DCs mature LPS blood-derived cells24_F_DCs_mat_CK DCs mature CK blood-derived cells 25_L_DCs + T DCs + Tcells blood-derived cells 26_Lym1 13987A1 Lymph Node Lymphoma 27_Lym243594B1 Muscle lymphoma 28_Lym3 65493A1 Testis Lymphoma 29_MalLym375894A1 Brain Lymphoma 30_NonHod_SCLym 83325A1 Lymph Node NHL Small Cell31_NonHod_FolLym 76943A1(5 tubes) Lymph Node NHL Follicular32_Lym_Fol_GI CN_4_ASRBNA35 NHL Follicular Grade I (Small Cell)33_Lym_Fol_GII CN_1_113GHA8J NHL Follicular Grade II (mixed Small &Large Cell) 34_Lym_Fol_GIII CN_8_VXML6AXI NHL Follicular Grade III(Large Cell) 35_MalLym1 76218B1 Testis NHL Large Cell 36_MalLym2 76102A1Lymph Node NHL Large Cell 37_Lym_DifBCell1 CN_2_4HDLNA2R NHL DiffuseLarge B-Cell 38_Lym_DifBCell2 CN_3_4M4S7AAM NHL Diffuse Large B-Cell39_Lym_DifBCell3 CN_5_HEODOAR2 NHL Diffuse Large B-Cell 40_NonHod_Lym177332A1(5 tubes) Colon NHL Diffuse Large B-Cell 41_MalLym4 76161A1Spleen NHL Diffuse Large B-Cell 42_Lym_MantleCell1 CN_6_MAE47AOY NHLMantle Cell 43_Lym_MantleCell2 CN_7_VJU9OAO9 NHL Mantle Cell44_NonHod_Lym2 95377A1(5 tubes) Spleen NHL 45_THP_1 THP-1 monocytes AMLcell line 46_KG_1 KG-1 myeloblast AML cell line 47_BDCM BDCM B and DClike AML cell line 48_CESS CESS lymphoblasts AML cell line 49_HL60 HL60myeloblast AML cell line 50_K562 K562 lymphoblasts CML cell line51_Jurkat Jurkat T lymphoblasts T ALL cell line 52_GA10 GA10 Blymphoblasts Burkitts lymphoma cell line 53_RAMOS RAMOS B lymphoblastsBurkitts lymphoma cell line 54_RAJI RAJI B lymphoblasts Burkittslymphoma cell line 55_Daudi Daudi B lymphoblasts Burkitts lymphoma cellline 56_NL564 -NL564 B lymphoblasts EBV transformed cell line 57_NL553NL553 B lymphoblasts EBV transformed cell line 58_SKW6.4 SKW6.4 B cellslymphoblasts EBV transformed cell line 59_NCI_H929 NCI-H929 Blymphoblasts Multiple Myeloma cell line 60_MC/CAR MC/CAR B lymphoblastsMultiple Myeloma cell line 61_U266 U266 B lymphoblasts Multiple Myelomacell line 62_RPMI8226 RPMI8226 B lymphoblasts Multiple Myeloma cell line63_IM_9 IM-9 B lymphoblasts Multiple Myeloma cell line 64_cereNcerebellum normal cerebellum normal 65_kidneyN1 kidney normal kidneynormal 66_kidneyN2 kidney normal kidney normal 67_KidneyN3 kidney normalkidney normal 68_colonN1 colon normal colon normal 69_colonN2 colonnormal colon normal 70_stomN stomach normal stomach normal 71_liverNliver normal liver normal 72_lungN1 lung normal lung normal 73_lungN2lung normal lung normal 74_small intestineN small intestine smallintestine 75_brainN brain normal mix brain normal mix 76_heartN heartnormal mix heart normal mix

TABLE 2 Tissue samples in normal panel: Sample id Sample id (GCI)/caseid Tissue id (Asterand)/ (Asterand) (GCI)/Specimen RNA id sample nameSource Lot no. id (Asternd) (GCI) 1—(7)-Bc-Rectum Biochain A6102972—(8)-Bc-Rectum Biochain A610298 3—GC-Colon GCI CDSUV CDSUVNR34—As-Colon Asterand 16364 31802 31802B1 5—As-Colon Asterand 22900 7444674446B1 6—GC-Small bowel GCI V9L7D V9L7DN6Z 7—GC-Small bowel GCI M3GVTM3GVTN5R 8—GC-Small bowel GCI 196S2 196S2AJN 9—(9)-Am-Stomach Ambion110P04A 10—(10)-Bc-Stomach Biochain A501159 11—(11)-Bc-Esoph BiochainA603814 12—(12)-Bc-Esoph Biochain A603813 13—As-Panc Asterand 8918 94429442C1 14—As-Panc Asterand 10082 11134 11134B1 16—As-Liver Asterand 79167203 7203B1 17—(28)-Am-Bladder Ambion 071P02C 18—(29)-Bc-BladderBiochain A504088 19—(64)-Am-Kidney Ambion 111P0101B 20—(65)-Cl-KidneyClontech 1110970 21—(66)-Bc-Kidney Biochain A411080 22—GC-Kidney GCIN1EVZ N1EVZN91 23—GC-Kidney GCI BMI6W BMI6WN9F 25—(43)-Bc-AdrenalBiochain A610374 26—(16)-Am-Lung Ambion 111P0103A 28—As-Lung Asterand9078 9275 9275B1 29—As-Lung Asterand 6692 6161 6161A1 30—As-LungAsterand 7900 7180 7180F1 31—(75)-GC-Ovary GCI L629FRV1 32—(76)-GC-OvaryGCI DWHTZRQX 33—(77)-GC-Ovary GCI FDPL9NJ6 34—(78)-GC-Ovary GCI GWXUZN5M36—GC-cervix GCI E2P2N E2P2NAP4 38—(26)-Bc-Uterus Biochain A50409039—(30)-Am-Placen Ambion 021P33A 40—(32)-Bc-Placen Biochain A41107341—GC-Breast GCI DHLR1 42—GC-Breast GCI TG6J6 43—GC-Breast GCI E6UDDE6UDDNCF 44—(38)-Am-Prostate Ambion 25955 45—Bc-Prostate BiochainA609258 46—As-Testis Asterand 13071 19567 19567B1 47—As-Testis Asterand19671 42120 42120A1 49—GC-Artery GCI YGTVY YGTVYAIN 50—TH-Blood-PBMCTel- 52497 Hashomer 51—TH-Blood-PBMC Tel- 31055 Hashomer52—TH-Blood-PBMC Tel- 31058 Hashomer 53—(54)-Ic-Spleen Ichilov CG-26754—(55)-Am-Spleen Ambion 111P0106B 54—(55)-Am-Spleen Ambion56—(58)-Am-Thymus Ambion 101P0101A 57—(60)-Bc-Thyroid Biochain A61028758—(62)-Ic-Thyroid Ichilov CG-119-2 59—Gc-Sali gland GCI NNSMV NNSMVNJC60—(67)-Ic-Cerebellum Ichilov CG-183-5 61—(68)-Ic-Cerebellum IchilovCG-212-5 62—(69)-Bc-Brain Biochain A411322 63—(71)-Bc-Brain BiochainA411079 64—(72)-Ic-Brain Ichilov CG-151-1 65—(44)-Bc-Heart BiochainA411077 66—(46)-Ic-Heart Ichilov CG-227-1 67—(45)-Ic-Heart IchilovCG-255-9 (Fibrotic) 68—GC-Skel Mus GCI T8YZS T8YZSN7O 69—GC-Skel Mus GCIQ3WKA Q3WKANCJ 70—As-Skel Mus Asterand 8774 8235 8235G1 71—As-Skel MusAsterand 8775 8244 8244A1 72—As-Skel Mus Asterand 10937 12648 12648C173—As-Skel Mus Asterand 6692 6166 6166A1

TABLE 3 Lung cancer testing panel sample id (GCI)/ case id TISSUE RNA(Asterand)/ ID ID lot (GCI)/ (GCI)/ no. specimen Sample Source/ sample(old ID ID Diag Specimen Tissue Delivery name samples) (Asterand)(Asterand) Diag remarks location Gr TNM LC GCI 1- 7Z9V4 7Z9V4AYM Aden BCGC- BAC- SIA LC GCI 2- ZW2AQ ZW2AQARP Aden BC GC- BAC- SIB LC Bioch 72-A501123 AC 2 UN (44)- Bc- BAC LC GCI 4- 3MOPL 3MOPLA79 Aden GC- Adeno-SIA LC GCI 5- KOJXD KOJXDAV4 Aden GC- Adeno- SIA LC GCI 6- X2Q44X2Q44A79 Aden GC- Adeno- SIA LC GCI 7- 6BACZ 6BACZAP5 Aden GC- Adeno-SIA LC GCI 8- BS9AF BS9AFA3E Aden GC- Adeno- SIA LC GCI 9- UCLOAUCLOAA9L Aden GC- Adeno- SIA LC GCI 10- BVYK3 BVYK3A7Z Aden GC- Adeno-SIA LC GCI 11- U4DM4 U4DM4AFZ Aden GC- Adeno- SIB LC GCI 12- OWX5YOWX5YA3S Aden GC- Adeno- SIB LC GCI 13- XYY96 XYY96A6B Aden GC- Adeno-SIIA LC GCI 14- SO7B1 SO7B1AIJ Aden GC- Adeno- SIIA LC GCI 15- QANSYQANSYACD Aden GC- Adeno- SIIIA LC Bioch 16- A610063 Aden 1 UN (95)- BC-Adeno LC Bioch 17- A609077 Aden 2-3 UN (89)- Bc- Adeno LC Bioch 18-A609218 Aden 3 UN (76)- Bc- Adeno LC Bioch 74- A504118 Aden 1 UN (2)-Bc- Adeno LC Bioch 75- A608301 Aden 2 UN (77)- Bc- Adeno LC Bioch 76-A609217 Aden 2 UN (75)- Bc- Adeno LC Bioch 78- A504116 Aden 2-3 UN (13)-Bc- Adeno LC Ichilov 81- CG- Aden UN UN (14)- 111 Ic- Adeno LC Aster 19-9220 9418 9418A1 SCC 1 TXN0M0 As- Sq- S0 LC GCI 20- U2QHS U2QHSA2N SCCGC- Sq- SIA LC GCI 21- TRQR7 TRQR7ACD SCC GC- Sq- SIB LC Aster 22- 1758132603 32603B1 SCC 3 T2N0M0 As- Sq- SIB LC Aster 23- 18309 41454 41454B1SCC 2 T2N0MX As- Sq- SIB LC Aster 24- 9217 9415 9415B1 SCC 2 T2N0M0 As-Sq- SIB LC GCI 25- RXQ1P RXQ1PAEA SCC GC- Sq- SIIB LC GCI 26- KB5KHKB5KHA6X SCC GC- Sq- SIIB LC GCI 27- LAYMB LAYMBALF SCC GC- Sq- SIIIA LCIchilov 29- CG- SCC UN UN (25)- 204 Ic-Sq LC Bioch 30- A408175 SCC 1 UN(19)- Bc- Sq LC Bioch 31- A607125 SCC 2 UN (78)- Bc- Sq LC Bioch 32-A409091 SCC 2 UN (16)- Bc- Sq LC Bioch 33- A609163 SCC 2 UN (80)- Bc- SqLC Bioch 34- A503387 SCC 2-3 UN (18)- Bc- Sq LC Bioch 35- A609076 SCC 3UN (81)- Bc- Sq LC Bioch 82- A503187 SCC 2 UN (21)- Bc- Sq LC Bioch 83-A503183 SCC 2 UN (17)- Bc- Sq LC Bioch 84- A609018 SCC 3 UN (79)- Bc- SqLC Bioch 85- A503386 SCC UN UN (22)- Bc- Sq LC Bioch 86- A501121 SCC UNUN (20)- Bc- Sq LC Bioch 87- A609219 SCC UN UN (88)- Bc- Sq LC Bioch 88-A409017 SCC UN UN (100)- Bc- Sq LC Ichilov 89- CG- SCC UN UN (24)- 123Ic-Sq LC GCI 36- AF8AL AF8ALAAL LCC GC- LCC- SIA LC GCI 37- O62XUO62XUA1X LCC GC- LCC- SIB LC GCI 38- OLOIM OLOIMAS1 LCC GC- LCC- SIB LCGCI 39- 1ZWSV 1ZWSVAB9 LCC GC- LCC- SIIB LC GCI 40- 2YHOD 2YHODA1H LCCNSCC . . . GC- LCC- SIIB LC GCI 41- 38B4D 38B4DAQK LCC GC- LCC- SIIB LCBioch 90- A504114 LCC UN UN (39)- Bc- LCC LC Bioch 91- A609165 LCC 3 UN(87)- Bc- LCC LC Bioch 92- A504113 LCC UN UN (38)- Bc- LCC LC Bioch 93-A609170 LCNC UN UN (82)- Bc- LCC LC GCI 42- QPJQL QPJQLAF6 SMCC NC 3 GC-SCC- SIB LC Bioch 43- A501391 SMCC UN (32)- Bc- SCC LC Bioch 44- A501389SMCC 3 UN (30)- Bc- SCC LC Bioch 45- A609162 SMCC UN UN (83)- Bc- SCC LCBioch 46- A608032 SMCC 3 UN (86)- Bc- SCC LC Bioch 47- A501390 SMCC UN(31)- Bc- SCC LC Bioch 48- A609167 SMCC UN UN (84)- Bc- SCC LC Bioch 49-A609169 SMCC UN UN (85)- Bc- SCC LC Bioch 50- A504115 SMCC UN (33)- Bc-SCC LN Aster 51- 9078 9275 9275B1 Norm-L PS As- N-PS LN Aster 52- 87578100 8100B1 Norm-L PM (Right), As- Lo N- be PM Inferior LN Aster 53-6692 6161 6161A1 Norm-L PM As- N- PM LN Aster 54- 7900 7180 7180F1Norm-L PM As- N- PM LN Aster 55- 8771 8163 8163A1 Norm-L PM (Left), As-Lo N- be PM Superior LN Aster 56- 13094 19763 19763A1 Norm-L PM As- N-PM LN Aster 57- 19174 40654 40654A2 Norm-L PM As- N- PM LN Aster 58-13128 19642 19642A1 Norm-L PM As- N- PM LN Aster 59- 14374 20548 20548C1Norm-L PM (Right), As- Lo N- be PM Superior LN Amb 60- 36856 N- PM (99)-PM Am-NPM LN Amb 61- 36853 N- PM (96)- PM Am-NPM LN Amb 62- 36854 N- PM(97)- PM Am-NPM LN Amb 63- 111P0103A N- PM- (93)- PM ICH Am-NPM LN Amb64- 36855 N- PM (98)- Am-NPM LN Bioch 69- A607257 N- PM (91)- P2- Bc-NPMPM LN Bioch 70- A608152 N- PM (90)- P2 Bc-NPM PM # of # Y. Cig. Use # Y.Source/ sample Smoking Per of off Tissue Delivery name CS Tum % Gen ageEthnic B Status day Tobacco Tobacco SmPY? LC GCI 1- IA 80 F 63 WCAU Prev20 15 27 N GC- U. BAC- SIA LC GCI 2- IB 70 F 56 WCAU Prev 15 28 10 Y GC-U. BAC- SIB LC Bioch 72- F 61 (44)- Bc- BAC LC GCI 4- IA 60 M 68 WCAUNev — — — N GC- U. Adeno- SIA LC GCI 5- IA 90 F 64 WCAU Prev 15 40  7 YGC- U. Adeno- SIA LC GCI 6- IA 85 M 58 WCAU Prev 10 47  0 Y GC- U.Adeno- SIA LC GCI 7- IA 60 F 65 WCAU Curr  6 30 — Y GC- U. Adeno- SIA LCGCI 8- IA 55 F 59 WCAU Curr 20 40 — N GC- U. Adeno- SIA LC GCI 9- IA 80F 69 WCAU Curr 30 52 — Y GC- U. Adeno- SIA LC GCI 10- IA 60 F 60 WCAUCurr 40 40 — N GC- U. Adeno- SIA LC GCI 11- IB 65 F 68 WCAU Prev  5  443 N GC- U. Adeno- SIB LC GCI 12- IB 90 M 69 WCAU Curr 10 — — GC- U.Adeno- SIB LC GCI 13- IIA 70 F 62 WCAU Prev  6 40  6 N GC- U. Adeno-SIIA LC GCI 14- IIA 70 M 56 WCAU Curr 30 25 — Y GC- U. Adeno- SIIA LCGCI 15- IIIA 65 F 61 WCAU Curr 30 36 — Y GC- U. Adeno- SIIIA LC Bioch16- F 54 (95)- BC- Adeno LC Bioch 17- M 62 (89)- Bc- Adeno LC Bioch 18-M 57 (76)- Bc- Adeno LC Bioch 74- M 64 (2)- Bc- Adeno LC Bioch 75- M 44(77)- Bc- Adeno LC Bioch 76- M 65 (75)- Bc- Adeno LC Bioch 78- M 64(13)- Bc- Adeno LC Ichilov 81- M 68 (14)- Ic- Adeno LC Aster 19- Occult80 M 67 CAU Curr 11-20 31-40 As- U. Sq- S0 LC GCI 20- IA 55 F 68 WCAUPrev 10 20  0 N GC- U. Sq- SIA LC GCI 21- IB 75 M 62 WCAU Prev 20 50  0Y GC- U. Sq- SIB LC Aster 22- IB 90 M 73 CAU Prev As- U. Sq- SIB LCAster 23- IB 100 M 66 CAU Prev 11-20 45 As- U. Sq- SIB LC Aster 24- IB90 M 65 CAU Curr  6-10 41-50 As- U. Sq- SIB LC GCI 25- IIB 55 F 44 WCAUPrev 20 20  0 Y GC- U. Sq- SIIB LC GCI 26- IIB 65 M 68 WCAU Prev 40 40 0 Y GC- U. Sq- SIIB LC GCI 27- IIIA 65 F 58 WCAU Prev 50 40  1 Y GC- U.Sq- SIIIA LC Ichilov 29- M 72 (25)- Ic-Sq LC Bioch 30- M 78 (19)- Bc- SqLC Bioch 31- M 62 (78)- Bc- Sq LC Bioch 32- F 68 (16)- Bc- Sq LC Bioch33- M 74 (80)- Bc- Sq LC Bioch 34- M 63 (18)- Bc- Sq LC Bioch 35- M 53(81)- Bc- Sq LC Bioch 82- M 52 (21)- Bc- Sq LC Bioch 83- M 57 (17)- Bc-Sq LC Bioch 84- M 67 (79)- Bc- Sq LC Bioch 85- M 48 (22)- Bc- Sq LCBioch 86- M 64 (20)- Bc- Sq LC Bioch 87- M 64 (88)- Bc- Sq LC Bioch 88-M 64 (100)- Bc- Sq LC Ichilov 89- M 76 (24)- Ic-Sq LC GCI 36- IA 85 M 45WCAU Prev 45 33  0 Y GC- U. LCC- SIA LC GCI 37- IB 75 F 60 WCAU Prev 3045  0 Y GC- U. LCC- SIB LC GCI 38- IB 70 M 68 WCAU Prev — 55 — Y GC- U.LCC- SIB LC GCI 39- IIB 50 M 51 WCAU Prev 20 12 22 Y GC- U. LCC- SIIB LCGCI 40- IIB 95 M 62 WCAU Prev 40 40  0 Y GC- U. LCC- SIIB LC GCI 41- IIB90 F 70 WCAU Prev 30 50 — Y GC- U. LCC- SIIB LC Bioch 90- F 35 (39)- Bc-LCC LC Bioch 91- F 47 (87)- Bc- LCC LC Bioch 92- M 58 (38)- Bc- LCC LCBioch 93- M 68 (82)- Bc- LCC LC GCI 42- IB 65 F 62 WCAU Prev 20 35   0.15 Y GC- U. SCC- SIB LC Bioch 43- M 30 (32)- Bc- SCC LC Bioch 44- M34 (30)- Bc- SCC LC Bioch 45- F 47 (83)- Bc- SCC LC Bioch 46- F 52 (86)-Bc- SCC LC Bioch 47- F 59 (31)- Bc- SCC LC Bioch 48- F 59 (84)- Bc- SCCLC Bioch 49- M 66 (85)- Bc- SCC LC Bioch 50- M (33)- Bc- SCC LN Aster51- M 22 CAU Nev As- U. N-PS LN Aster 52- F 26 CAU Nev As- U. N- PM LNAster 53- M 37 CAU Nev As- U. N- PM LN Aster 54- F 76 CAU Prev As- U. N-PM LN Aster 55- M 81 CAU Prev 41 31-40 As- U. or N- more PM LN Aster 56-M 0 CAU Prev 21-40 41-50 As- U. N- PM LN Aster 57- F 69 CAU Curr 21-4031-40 As- U. N- PM LN Aster 58- F 75 CAU As- N- PM LN Aster 59- F 75 CAUAs- N- PM LN Amb 60- M 31 (99)- Am-NPM LN Amb 61- F 43 (96)- Am-NPM LNAmb 62- M 46 (97)- Am-NPM LN Amb 63- F 61 (93)- Am-NPM LN Amb 64- F 72(98)- Am-NPM LN Bioch 69- P2 24, (91)- 29 Bc-NPM LN Bioch 70- P2 27,(90)- 28 Bc-NPM Cause Source/ sample # Recovery of Exc. Tissue Deliveryname Smppl DrAl Dr Type Death Y. LC GCI 1- — Y  0 Surg 2001 GC- BAC- SIALC GCI 2- 1 Y  6 Surg 2002 GC- BAC- SIB LC Bioch 72- (44)- Bc- BAC LCGCI 4- — N — Surg 2001 GC- Adeno- SIA LC GCI 5- 1 N  0 Surg 2003 GC-Adeno- SIA LC GCI 6- 2 N — Surg 2004 GC- Adeno- SIA LC GCI 7- 1 N — Surg2004 GC- Adeno- SIA LC GCI 8- — N — Surg 2004 GC- Adeno- SIA LC GCI 9- 4N — Surg 2005 GC- Adeno- SIA LC GCI 10- — N — Surg 2002 GC- Adeno- SIALC GCI 11- — N — Surg 2003 GC- Adeno- SIB LC GCI 12- — N — Surg 2002 GC-Adeno- SIB LC GCI 13- — Y 0 Surg 2004 GC- Adeno- SIIA LC GCI 14- 1 N —Surg 2001 GC- Adeno- SIIA LC GCI 15- 1 N — Surg 2004 GC- Adeno- SIIIA LCBioch 16- (95)- BC- Adeno LC Bioch 17- (89)- Bc- Adeno LC Bioch 18-(76)- Bc- Adeno LC Bioch 74- (2)- Bc- Adeno LC Bioch 75- (77)- Bc- AdenoLC Bioch 76- (75)- Bc- Adeno LC Bioch 78- (13)- Bc- Adeno LC Ichilov 81-(14)- Ic- Adeno LC Aster 19- O Surg 2003 As- Sq- S0 LC GCI 20- — N —Surg 2004 GC- Sq- SIA LC GCI 21- 5 N — Surg 2005 GC- Sq- SIB LC Aster22- O Surg 2004 As- Sq- SIB LC Aster 23- P Surg 2005 As- Sq- SIB LCAster 24- O Surg 2002 As- Sq- SIB LC GCI 25- 2 N — Surg 2004 GC- Sq-SIIB LC GCI 26- 2 N — Surg 2004 GC- Sq- SIIB LC GCI 27- 2 N — Surg 2004GC- Sq- SIIIA LC Ichilov 29- (25)- Ic-Sq LC Bioch 30- (19)- Bc- Sq LCBioch 31- (78)- Bc- Sq LC Bioch 32- (16)- Bc- Sq LC Bioch 33- (80)- Bc-Sq LC Bioch 34- (18)- Bc- Sq LC Bioch 35- (81)- Bc- Sq LC Bioch 82-(21)- Bc- Sq LC Bioch 83- (17)- Bc- Sq LC Bioch 84- (79)- Bc- Sq LCBioch 85- (22)- Bc- Sq LC Bioch 86- (20)- Bc- Sq LC Bioch 87- (88)- Bc-Sq LC Bioch 88- (100)- Bc- Sq LC Ichilov 89- (24)- Ic-Sq LC GCI 36- 2 Y28 Surg 2004 GC- LCC- SIA LC GCI 37- 3 N — Surg 2004 GC- LCC- SIB LC GCI38- — N — Surg 2001 GC- LCC- SIB LC GCI 39- 1 N — Surg 2004 GC- LCC-SIIB LC GCI 40- 2 Y 12 Surg 2004 GC- LCC- SIIB LC GCI 41- 2 Y 13 Surg2002 GC- LCC- SIIB LC Bioch 90- (39)- Bc- LCC LC Bioch 91- (87)- Bc- LCCLC Bioch 92- (38)- Bc- LCC LC Bioch 93- (82)- Bc- LCC LC GCI 42- 2 N —Surg 2003 GC- SCC- SIB LC Bioch 43- (32)- Bc- SCC LC Bioch 44- (30)- Bc-SCC LC Bioch 45- (83)- Bc- SCC LC Bioch 46- (86)- Bc- SCC LC Bioch 47-(31)- Bc- SCC LC Bioch 48- (84)- Bc- SCC LC Bioch 49- (85)- Bc- SCC LCBioch 50- (33)- Bc- SCC LN Aster 51- NU Surg 2003 As- N-PS LN Aster 52-O Aut CA 2003 As- N- PM LN Aster 53- C Aut MCE 2002 As- N- PM LN Aster54- Aut CPulA 2002 As- N- PM LN Aster 55- O Aut CA 2003 As- N- PM LNAster 56- P Aut IC As- N- PM LN Aster 57- P Aut CPulA 2005 As- N- PM LNAster 58- Aut CPulA 2004 As- N- PM LN Aster 59- Aut CerA 2004 As- N- PMLN Amb 60- (99)- Am-NPM LN Amb 61- (96)- Am-NPM LN Amb 62- (97)- Am-NPMLN Amb 63- (93)- Am-NPM LN Amb 64- Am-NPM LN Bioch 69- (91)- Bc-NPM LNBioch 70- (90)- Bc-NPM

TABLE 3_1 Key Full Name # Cig. Per day Number of Cigarettes per day # DrNumber of Drinks # of Y. Use of Number of Years Using Tobacco Tobacco #Y. off Tobacco Number of Years Off Tobacco AC Alveolus carcinoma AdenADENOCARCINOMA Amb Ambion Aster Asterand Aut Autopsy BCBRONCHIOLOALVEOLAR CARCINOMA Bioch Biochain C Current Use CA Cardiacarrest CAU Caucasian Cer A Cerebrovascular accident CPul ACardiopulmonary arrest CS Cancer Stage Curr U. Current Use DiagDiagnosis Dr Al Drink Alcohol? Exc Y. Excision Year Gen Gender Gr GradeHeight HT IC Ischemic cardiomyopathy LC Lung Cancer LCC LARGE CELLCARCINOMA LCNC Large Cell Neuroendocrine Carcinoma LN Lung Normal MCEMassive cerebral edema N No NC NEUROENDOCRINE CARCINOMA Nev. U. NeverUsed Norm-L Normal Lung N-P2-PM Normal (Pool 2)- PM N-PM Normal-PM NSCC. . . NON-SMALL CELL CARCINOMA WITH SARCOMUTOUS TRANSFORMTAIO NU Neverused O Occasional Use P Previous Use P2 Pool 2 Prev U. Previous Use SCCSquamous Cell Carcinoma Sm P Y? Have people at home smoked in past 15 yrSm ppl If yes, how many? SMCC SMALL CELL CARCINOMA SMOKE_GROWING_UP Didpeople smoke at home while growing up Surg Surgical Tum % TumorPercentage WCAU White Caucasian Y Yes

TABLE 4 Tissue samples in ovary panel sample_id (GCI)/ case id RNA(Asterand)/ ID lot (GCI)/ Age no. Sample Meno- at Oral Oral Source/sample (old ID C Tumor Ethnic pausal Mens Preg Preg first Con Con TubalRecovery Tissue Delivery name samples) (Asterand) Diag Stage % age BGCA125PRE Status Age Times Toterm child OC Length Unit ligation Type OVCAsterand 1- 23074 71900A2 SA I 80 49 CAU Pre-M 2 1 Surg As- SerSI OVCAsterand 2- 22653 70270A1 SA I 90 69 WCAU Post-M 1 1 Surg As- SerSI OVCAsterand 3- 18700 40771B1 SA IB 100 62 WCAU Post-M 3 3 Surg As- SerSIBOVC GOG 79- 93-09- SPC 1B 67 (32)- 4901 GO- SerSIB OVC Asterand 4- 1764632667B1 SA IB 100 68 W Post-M 9 2 Surg As- SerSIB OVC Asterand 5- 1564422996A1 SA IC 100 48 CAU M 4 2 Surg As- SerSIC OVC Asterand 6- 1870140773C1 SA IIA 100 59 CAU Post-M 1 1 Surg As- SerSIIA OVC GCI 7- 2O37OSA IIB 75 43 WCAU — Pre-M 12 0 0  0 NO — NO Surg GC- SerSIIB OVC GCI 8-7B3DP SA IIB 70 70 WCAU — Post-M 14 5 3 20 YES 6 months NO Surg GC-SerSIIB OVC GOG 80- 2001- PSC 3A 72 (30)- 08- GO- G011 SerSIIIA OVC GOG81- 95-08- PSA 3B 50 (70)- G069 GO- SerSIIIB OVC GOG 82- 99-12- A 3C46 >500 (5)- G432 GO- SerSIIIC OVC Asterand 9- 13268 19832A1 SA IIIC 9048 C Post-M Surg As- SerSIIIC OVC GOG 83- 2001- SA 3C 50 260 (29)- 12-GO- G035 SerSIIIC OVC GCI 10- 3NTIS SA IIIC 70 53 WCAU 70 Post-M 12 1 126 YES 3 months NO Surg GC- SerSIIIC OVC GCI 11- CEJUS SA IIIC 70 53WCAU 4814 Pre-M — 2 2 30 NO — NO Surg GC- SerSIIIC OVC GCI 12- 5NCLK SAIIIC 70 54 WCAU 209 Post-M 13 2 2 21 YES 1 years NO Surg GC- SerSIIICOVC ABS 84- N0021 PSA 3C 55 CAU (25)- AB- SerSIIIC OVC GCI 13- 1HI5H SAIIIC 90 61 WCAU 34 Post-M 12 6 3 22 NO — NO Surg GC- SerSIIIC OVC GCI14- 7RMHZ SA IIIC 80 63 WCAU — Post-M 12 2 2 20 YES 10  years NO SurgGC- SerSIIIC OVC GCI 15- 4WAAB SA IIIC 90 63 WCAU — Post-M 11 2 1 29 YES4 years NO Surg GC- SerSIIIC OVC GCI 16- 79Z67 SA IIIC 85 67 WCAU —Post-M 12 6 5 24 YES 2 years YES Surg GC- SerSIIIC OVC GOG 85- 94-05-APP 3C 67 (13)- 7603 GO- SerSIIIC OVC GCI 17- DDSNL SA IIIC 70 68 WCAU —Post-M 11 4 4 19 NO — NO Surg GC- SerSIIIC OVC GCI 18- DH8PH SA IV 95 70WCAU — Post-M 13 4 3 20 NO — NO Surg GC- SerSIV OVC BioChain 86- A503175SPC 41 Asian (33)- BC- Ser OVC BioChain 87- A501111 A 41 Asian (14)- Bc-Ser OVC Biochain 88- A406023 A 45 Asian (12)- Bc- Ser OVC Biochain 89-A407068 A 49 Asian (11)- Bc- Ser OVC ABS 90- ILS- PC UN 50 Asian (4)-7286 AB- Ser OVC ABS 91- A0106 A UN 51 Asian (6)- AB- Ser OVC ABS 92-ILS- PA UN 52 Asian (3)- 1431 AB- Ser OVC BioChain 93- A503176 SPC 52Asian (31)- Bc- Ser OVC ABS 94- ILS- PA UN 53 Asian (2)- 1408 AB- SerOVC ABS 95- IND- A 59 Asian (7)- 00375 AB- Ser OVC BioChain 96- A501113A 60 Asian (8)- Bc- Ser OVC Biochain 97- A407069 A 60 Asian (10)- Bc-Ser OVC ABS 98- ILS- PA UN 73 Asian (1)- 1406 AB- Ser OVC GCI 19- E2WKFEA IA 70 30 WCAU — Pre-M 12 6 5 17 YES 6 years NO Surg GC- EndoSIA OVCGCI 20- 5895C EA IA 95 39 WCAU — Pre-M 14 2 2 20 NO — NO Surg GC-EndoSIA OVC GCI 21- 533DX EA IA 95 50 WCAU 190 Pre-M 11 0 — — YES 2years NO Surg GC- EndoSIA OVC GCI 22- HZ2EY EA IA 90 55 WCAU 1078 Pre-M13 0 — — NO — NO Surg GC- EndoSIA OVC GCI 23- RWOIV EA IA 65 47 WCAU1695 Pre-M 14 0 — — NO — NO Surg GC- EndoSIA OVC GCI 24- 1U52X EA IIA 9561 WCAU 275 — — — — — Surg GC- EndoSIIA OVC GCI 25- A17WS EA IIB 70 67WCAU 78 Post-M 14 0 — — NO — NO Surg GC- EndoSIIB OVC GCI 26- 1VT3I EAIIIC 90 50 WCAU — Pre-M 12 2 2 24 YES 1 years NO Surg GC- EndoSIIIC OVCGCI 27- PZQXH EA IIIC 80 52 WCAU — Pre-M 11 0 — — YES 5 years NO SurgGC- EndoSIIIC OVC GCI 28- I8VHZ EA IV 90 68 WCAU — Post-M — 2 2 27 NO —NO Surg GC- EndoSIV OVC GOG 99- 98-03- Mixed . . . 2 38 >35 (41)- G803GO- SerMixSII OVC GOG 100- 95-11- PS & 3C 49 (40)- G006 EC GO-SerMixSIIIC OVC GOG 101- 2002- MS & 3C 56 (37)- 05- EA GO- G513SerMixSIIIC OVC GOG 102- 2002- MS & 3C 64 (38)- 05- EAM GO- G509SerMixSIIIC OVC GOG 103- 95-04- PEA 3C 68 (34)- 2002 GO- SerMixSIIIC OVCGOG 29- 95-10- MC IA 44 >100 (21)- G020 GO- MucSIA OVC GCI 30- IMDA1 MAIC 70 41 WCAU 50 Pre-M 12 2 1 24 NO — Surg GC- MucSIC OVC Asterand 31-12742 18920A1 MA IC 70 61 C Post-M 3 3 Surg As- MucSIC OVC ABS 32- A0139MC IC 72 Asian (22)- AB- MucSIC OVC GCI 33- NJM4U MA IIA 80 51 WCAU SurgGC- MucSIIA OVC ABS 34- USA- PMC IIIA 45 C (20)- 00273 AB- MucSIIIA OVCGCI 35- RAFCW MA IIIA 75 55 WCAU 95 Post-M 13 4 3 22 NO — NO Surg GC-MucSIIIA OVC Asterand 36- 23177 72888A1 MA IIIC 60 52 C Pre-M Surg As-MucSIIIC OVC Asterand 37- 16103 29374B1 MA IIIC 100 62 W Post-M 1 1 SurgAs- MucSIIIC OVC BioChain 104- A504085 MA 34 Asian (19)- Bc- Muc OVCBioChain 105- A504083 MA 45 Asian (18)- Bc- Muc OVC BioChain 106-A504084 MA 51 Asian (17)- Bc- Muc OVC BioChain 107- A407065 C 27 Asian(15)- Bc- Car OVC Clontech 108- 1090387 CNOS 58 Asian (16)- Cl- Car OVCGOG 109- 2001- CCA 1A 73 (44)- 07- GO- G084 ClearcellSIA OVC GOG 110-2001- CCA 3A 74 slightly (43)- 10- elevated GO- G002 ClearcellSIIIAOVC_BT GCI 38- SC656 MBT IA 75 40 WCAU 138 Pre-M 13 2 2 23 NO — YES SurgGC- MucBorderSIA OVC_BT GCI 39- 3D5FO MBT IA 85 51 WCAU 19 ? 15 0 — — NO— NO Surg GC- MucBorderSIA OVC_BT GCI 40- 7JP3F MBT IA 75 56 WCAU 125Post-M 14 3 3 19 YES 5 years NO Surg GC- MucBorderSIA OVC_BT ABS 111-VNM- MCLowM 45 Asian (23)- 00187 AB- Border OVC_BT GOG 112- 98-08- EA 1A46 (42)- G001 of GO- BM BorderSIA OVC_B GOG 41- 99-10- BMC 32 6 (62)-G442 Go- BenMuc OVC_B GCI 43- QLIKY BMC 100 42 WCAU Surg GC- BenMucOVC_B Asterand 44- 16870 30534A1 BMC 100 45 W Pre-M 2 2 Surg As- BenMucOVC_B GOG 45- 99-01- BMC 46 (56)- G407 GO- BenMuc OVC_B GCI 46- 943ECBMC 75 54 WCAU Surg GC- BenMuc OVC_B GCI 47- JO8W7 BMC 50 56 WCAU SurgGC- BenMuc OVC_B Asterand 48- 17016 30645B1 BSC IA 100 38 C Pre-M 2 2Surg As- BenSer OVC_B GOG 49- 99-06- BSC 57 (64)- G039 GO- BenSer OVC_BGCI 50- DQQ2F BSCF 95 68 WCAU Surg GC- BenSer OVC_B Asterand 51- 87868275A1 BSC 100 80 CAU Post-M 10  9 Surg As- BenSer OVC_NBM Asterand 52-15690 23054A1 NO- 52 CAU Pre-M 10  3 Surg As-NBM BM OVC_NBM Asterand 53-16843 30488A1 NO- 57 W Post-M 4 2 Surg As-NBM BM OVC_NBM Asterand 54-16850 30496B1 NO- 65 W Post-M 2 2 Surg As-NBM BM OVC_NBM Asterand 55-16848 30499C1 NO- 66 CAU Post-M 9 2 Surg As-NBM BM OVC_N GCI 56- WPU1UNO- 0 32 WC Surg GC-NPS PS OVC_N GCI 57- Y9VHI NO- 0 35 WCAU Surg GC-NPSPS OVC_N GCI 58- 76VM9 NO- 0 41 WCAU Surg GC-NPS PS OVC_N GCI 59- DWHTZNO- 0 42 WCAU Surg GC-NPS PS OVC_N GCI 60- SJ2R2 NO- 0 43 WCAU SurgGC-NPS PS OVC_N GCI 61- 9RQMN NO- 0 45 WCAU Surg GC-NPS PS OVC_N GCI 62-TOAE5 NO- 0 45 WCAU Surg GC-NPS PS OVC_N GCI 63- TW9PM NO- 0 46 WCAUSurg GC-NPS PS OVC_N GCI 64- 2VND2 NO- 0 46 WCAU Surg GC-NPS PS OVC_NGCI 65- L629F NO- 0 47 WCAU Surg GC-NPS PS OVC_N GCI 66- XLB23 NO- 0 47WCAU Surg GC-NPS PS OVC_N GCI 67- IDUVY NO- 0 47 WCAU Surg GC-NPS PSOVC_N GCI 68- ZCXAD NO- 0 48 WCAU Surg GC-NPS PS OVC_N GCI 69- PEQ6C NO-0 49 WCAU Surg GC-NPS PS OVC_N GCI 70- DD73B NO- 0 49 WCAU Surg GC-NPSPS OVC_N GCI 71- E2UF7 NO- 0 53 WCAU Surg GC-NPS PS OVC_N GCI 72- GWXUZNO- 0 53 WCAU Surg GC-NPS PS OVC_N GCI 73- 4YG5P NO- 0 55 WCAU SurgGC-NPS PS OVC_N GCI 74- FDPL9 NO- 0 56 WCAU Surg GC-NPS PS OVC_NBioChain 75- A503274 NO- 41 Asian (45)- PM Bc-NPM OVC_N BioChain 76-A504086 NO- 41 Asian (46)- PM Bc-NPM OVC_N Ichilov 77- CG- NO- 49 (71)-188-7 PM Ic-NPM OVC_N BioChain 78- A504087 NO- 51 Asian (48)- PM Bc-NPM

TABLE 4_1 Key Full Name A Adenocarcinoma APP Adenocarcinoma from primaryperitoneal BMC BENIGN MUCINOUS CYSTADENOMA BSC BENIGN SEROUS CYSTADENOMABSCF BENIGN SEROUS CYSTADENOFIBROMA C Carcinoma C Stage Cancer stage CAUCaucasian CCA Clear cell adenocarcinoma CNOS Carcinoma NOS EAENDOMETROID ADENOCARCINOMA EA of Endometroid adenocarcinoma ofborderline BM malignancy M Menopausal MA MUCINOUS ADENOCARCINOMA MBTMUCINOUS BORDERLINE TUMOR MC Mucinous cystadenocarcinoma MC Low Mucinouscystadenocarcinoma with low malignant M Mens. Menstrual Age Age Mixed .. . Mixed epithelial cystadenocarcinoma with mucinous, endometrioid,squamous and papillary serous MS & Mixed serous and endometrioidadenocarcinoma EA MS & Mixed serous and endometrioid adenocarcinoma ofEAM mullerian NO-BM NORMAL OVARY-BM NO-PM NORMAL OVARY-PM NO-PS NORMALOVARY-PS OC Oral Contraceptive OVC Ovary Cancer OVC_B Ovary BenignOVC_BT Ovary Borderline Tumor OVC_N Ovary Normal OVC_NBM Ovarynormal-benign matched PA Papillary adenocarcinoma PC Papillarycystadenocarcinoma PEA Papillary endometrioid adenocarcinoma PMCPapillary mucinous cystadenocarcinoma Post-M Post-menopausal Pre-MPre-menopausal PS & EC Papillary serous and endometrioidcystadenocarcinoma PSA Papillary serous adenocarcinoma PSC Papillaryserous carcinoma SA SEROUS ADENOCARCINOMA SPC Serous papillarycystadenocarcinoma W White WCAU WHITE/CAUCASIAN

TABLE 5 Colon cancer testing panel sample_id (GCI)/ case id TISSUE(Asterand)/ ID lot (GCI)/ no. specimen Sample Source/ sample (old ID IDDiag Specimen Tissue Delivery name samples (Asterand) (Asterand) Diagremarks location Gr TNM CS CC Asterand 1- 18036 31312 31312B1 Aden Cec 3TXN0M0 0 As- AdenS0 CC GCI 2- 4QDH8 4QDH8ADT Aden DisC I GC- AdenoSI CCIchilov 3- CG- AI Rectum UN I (7)- 235 Ic- AdenoSI CC GCI 4- NTAI8NTAI8AOU Aden Cec I GC- AdenoSI CC GCI 5- ARA7P ARA7PAQA Aden Ret, I GC-Low AdenoSI Ant CC Ichilov 6- CG- UA 3 IIA (20)- 249 Ic- AdenoSIIA CCGCI 7- AFTS6 AFTS6AP6 Aden IIA GC- AdenoSIIA CC GCI 8- 5CYDK 5CYDKACSAden IIA GC- AdenoSIIA CC GCI 9- XKSLS XKSLSAF7 Aden IIA GC- AdenoSIIACC GCI 10- B4RU8 B4RU8A8Q Aden IIA GC- AdenoSIIA CC GCI 11- HB8EYHB8EYA8I Aden IIA GC- AdenoSIIA CC Ichilov 12- CG- Aden 2 II (22)- 229CAdenoSII CC GCI 13- X8C7X X8C7XATL Aden IIA GC- AdenoSIIA CC GCI 14-HCP6K HCP6KA8Z Aden IIA GC- AdenoSIIA CC GCI 15- ZX4X7 ZX4X7AXA Aden IIAGC- AdenoSIIA CC Asterand 16- 17915 31176 31176A1 Aden 2-3 T3N0M0 IIAAs- AdenoSIIA CC Ichilov 17- CG- Aden Cec 2 IIA (1)- 335 Ic- AdenoSIIACC Asterand 19- 12772 18885 18885A1 Aden rectum 2 T3NXM0 IIA As-AdenoSIIA CC GCI 20- JFYXP JFYXPAMP Aden IIA GC- AdenoSIIA CC GCI 21-OJXW9 OJXW9ASR Aden IIA GC- AdenoSIIA CC Ichilov 22- CG- Aden sigma 2IIA (28)- 284 Ic- AdenoSIIA CC Ichilov 23- CG- Aden SigCol 1-2 eIIA(10)- 311 Ic- AdenoSIIA CC Ichilov 24- CG- WPAden Rectum III (14)- 222Ic- (2) AdenoSIII CC Ichilov 25- CG- MA sigma UN III (23)- 282 Ic-AdenoSIII CC GCI 26- OTPI7 OTPI7AWY Aden III GC- AdenoSIII CC GCI 27-IG9NK IG9NKAD3 MA III GC- AdenoSIII CC GCI 28- 53OM7 53OM7AGL Aden IIIGC- AdenoSIII CC GCI 29- BLUW6 BLUW6A6Y Aden III GC- AdenoSIII CC GCI30- VZ6QA VZ6QAAFA Aden RECTUM III GC- AdenoSIII CC Ichilov 31- CG- Aden1-2 III (6)- 303 Ic- (3) AdenoSIII CC Ichilov 32- CG- Aden Cecum 2 III(2)- 307 Ic- AdenoSIII CC Ichilov 33- CG- Aden 1-2 III (11)- 337 Ic-AdenoSIII CC Asterand 34- 18462 40971 40971A1 TA SigCol 2 TXN2M0 IIICAs- AdenoSIIIC CC Ichilov 35- CG- Aden RectCol 2 IV (13)- 290 Ic-AdenoSIV CC GCI 36- 7D7QV 7D7QVAE6 Aden IV GC- AdenoSIV CC GCI 37- 38U4V38U4VAA4 Aden IV GC- AdenoSIV CC Ichilov 38- CG- Aden Rectum 2 IV (9)-297 Ic- AdenoSIV CC Ichilov 71- CG- Aden 2 (16)- 278C Ic- Adeno CCIchilov 72- CG- Carc 3 (4)- 276 Ic- Adeno CC Ichilov 73- CG- Aden Rectum2 (17)- 163 Ic- Adeno CC Ichilov 74- CG- Aden ColSig 2 (5)- 308 Ic-Adeno CC Ichilov 75- CG- Aden 3 (72)- 309 Ic- Adeno CC Ichilov 76- CG-Aden UN (18)- 22C Ic- Adeno CC Ichilov 78- CG- Aden UN (21)- 18C Ic-Adeno CC Ichilov 79- CG-12 Aden UN (24)- Ic- Adeno CC Ichilov 80- CG-2Aden UN (25)- Ic- Adeno CC biochain 82- A606258 Aden, 2 (61)- Ulcer Bc-Adeno CC biochain 83- A609150 Aden 3 (57)- Bc- Adeno CC biochain 84-A609148 Aden 2 (56)- Bc- Adeno CC biochain 85- A609161 Aden 3 (53)- Bc-Adeno CC biochain 86- A609142 Aden 3 (54)- Bc- Adeno CC biochain 87-A609059 Aden, 1 (59)- Ulcer Bc- Adeno CC biochain 88- A609058 Aden, 2(60)- Ulcer Bc- Adeno CC biochain 89- A609144 Aden 3 (55)- Bc- Adeno CCbiochain 90- A609152 Aden 1 (58)- Bc- Adeno CB GCI 40- IG3OY IG3OYN7STSAden RTCol GC- Ben CB GCI 41- GKIEY GKIEYAV4 TSAdenHGD ProxTCol GC-Ben CN GCI 42- AGVTC AGVTCNK7 NC DIV GC-NPS CN Asterand 43- 8956 91539153B1 NC As-NPS CN GCI 44- IG3OY IG3OYN7S NC RTCol GC-NPS CN GCI 45-K9OYX K9OYXN4F NC Divsw/FDIV LTCol GC-NPS CN Asterand 46- 23024 7444574445B1 NC ChrDivs As-NPS CN Asterand 47- 23049 71410 71410B2 NC ChrDivsAs-NPS CN GCI 48- G7JJX G7JJXAX7 NC Divsw/DIV... SigCol GC-NPS CNAsterand 49- 22900 74446 74446B1 NC ADw/AF As-NPS CN GCI 50- XVPZ2XVPZ2NDD NC Div GC-NPS CN GCI 51- CDSUV CDSUVNR3 NC CU GC-NPS CN GCI 52-GP5KH GP5KHAOC NC Div GC-NPS CN GCI 53- YUZNR YUZNRNDN NC Divs SigColGC-NPS CN GCI 54- 28QN6 28QN6NI1 NC TSAden RTCol GC-NPS CN GCI 55- GV6N8GV6N8NG9 NC Divs, GC-NPS PA CN GCI 56- ZJ17R ZJ17RNIH NC TubAden RTColGC-NPS CN GCI 57- 2EEBJ 2EEBJN2Q NC Div/ GC-NPS ChrInfl CN GCI 58- 68IX568IX5N1H NC ChrDiv LTCol GC-NPS CN GCI 59- 9GEGL 9GEGLN1V NC ExtDivsSigCol GC-NPS CN GCI 60- PKU8O PKU8OAJ3 NC Divs, SigCol GC-NPS ChrDiv...CN Asterand 61- 22903 74452 74452B1 NC MUw/MI As-NPS CN Asterand 62-16364 31802 31802B1 NC UC As-NPS CN biochain 63- A607115 N- PM (65)- PMBc-NPM CN Ambion 64- 071P10B N- PM (71)- PM Am-NPM CN biochain 65-A609262 N- PM (66)- PM Bc-NPM CN biochain 66- A609260 N- PM (63)- PMBc-NPM CN biochain 67- A608273 N- PM (62)- PM Bc-NPM CN biochain 68-A609261 N- PM (64)- PM Bc-NPM CN biochain 69- A501156 N- PM (41)- PMBc-NPM CN biochain 70- A406029 + N- PM (67)- A411078 PMP10 Bc-NPM Dr.Source/ sample Alcohol per Alc. Recovery Exc. Tissue Delivery name CS2Tumor % Gender age Ethnic B Status day Dur. Type Y. CC Asterand 1- 80 F43 CAU NU Auto 2004 As- AdenS0 CC GCI 2- DukeA 85 F 44 WCAU Y 4 Surg GC-AdenoSI CC Ichilov 3- DukeA F 66 (7)- Ic- AdenoSI CC GCI 4- DukeB1 80 M53 WCAU Y — Surg GC- AdenoSI CC GCI 5- DukeB1 70 F 70 WCAU Y 0 Surg GC-AdenoSI CC Ichilov 6- DukeB2 M 36 (20)- Ic- AdenoSIIA CC GCI 7- DukeB275 M 39 WCAU N 0 Surg GC- AdenoSIIA CC GCI 8- DukeB2 65 M 44 WCAU N —Surg GC- AdenoSIIA CC GCI 9- DukeB2 65 M 48 WCAU Y 10  Surg GC-AdenoSIIA CC GCI 10- DukeB2 65 F 50 WCAU N — Surg GC- AdenoSIIA CC GCI11- DukeB2 65 M 53 WCAU N — Surg GC- AdenoSIIA CC Ichilov 12- DukeB F 55(22)- AdenoSII CC GCI 13- DukeB2 90 M 56 WCAU N — Surg GC- AdenoSIIA CCGCI 14- DukeB2. 80 M 58 WCAU Y 4 Surg GC- AdenoSIIA CC GCI 15- DukeB2 90M 60 WCAU Y 5 Surg GC- AdenoSIIA CC Asterand 16- DukeB2 60 F 64 CAU occ1 21-30 Auto 2004 As- drink/ years AdenoSIIA week CC Ichilov 17- DukeB2F 66 (1)- Ic- AdenoSIIA CC Asterand 19- DukeB2 60 F 67 CAU NU Surg 2004As- AdenoSIIA CC GCI 20- DukeB2 60 F 68 WCAU Y — Surg GC- AdenoSIIA CCGCI 21- DukeB2 90 F 69 WCAU N — Surg GC- AdenoSIIA CC Ichilov 22- DukeB2F 72 (28)- Ic- AdenoSIIA CC Ichilov 23- DukeB2 M 88 (10)- Ic- AdenoSIIACC Ichilov 24- DukeC F 49 (14)- Ic- AdenoSIII CC Ichilov 25- DukeC M 51(23)- Ic- AdenoSIII CC GCI 26- DukeC2 70 F 54 WCAU N — Surg GC-AdenoSIII CC GCI 27- DukeC2 90 F 54 WCAU N — Surg GC- AdenoSIII CC GCI28- DukeC2 75 F 61 WCAU N — Surg GC- AdenoSIII CC GCI 29- DukeC2 85 F 64WCAU N — Surg GC- AdenoSIII CC GCI 30- DukeC2 60 M 67 WCAU Y 14  SurgGC- AdenoSIII CC Ichilov 31- DukeC2. F 77 (6)- Ic- AdenoSIII CC Ichilov32- DukeC2. F 89 (2)- Ic- AdenoSIII CC Ichilov 33- DukeC2. NA NA (11)-Ic- AdenoSIII CC Asterand 34- 76 F 68 CAU NU Surg 2005 As- AdenoSIIIC CCIchilov 35- DukeD. M 47 (13)- Ic- AdenoSIV CC GCI 36- DukeD 80 F 52 WCAUY 3 Surg GC- AdenoSIV CC GCI 37- DukeD 85 F 53 WCAU — Surg GC- AdenoSIVCC Ichilov 38- DukeD. M 62 (9)- Ic- AdenoSIV CC Ichilov 71- UN 50 F 60(16)- Ic- Adeno CC Ichilov 72- UN 75 M 64 (4)- Ic- Adeno CC Ichilov 73-UN M 73 (17)- Ic- Adeno CC Ichilov 74- UN F 80 (5)- Ic- Adeno CC Ichilov75- UN F 88 (72)- Ic- Adeno CC Ichilov 76- UN NA NA (18)- Ic- Adeno CCIchilov 78- UN NA NA (21)- Ic- Adeno CC Ichilov 79- UN NA NA (24)- Ic-Adeno CC Ichilov 80- UN NA NA (25)- Ic- Adeno CC biochain 82- UN M 41(61)- Bc- Adeno CC biochain 83- UN F 45 (57)- Bc- Adeno CC biochain 84-UN 40 F 48 (56)- Bc- Adeno CC biochain 85- UN F 53 (53)- Bc- Adeno CCbiochain 86- UN M 53 (54)- Bc- Adeno CC biochain 87- UN M 58 (59)- Bc-Adeno CC biochain 88- UN M 67 (60)- Bc- Adeno CC biochain 89- UN M 68(55)- Bc- Adeno CC biochain 90- UN M 73 (58)- Bc- Adeno CB GCI 40- F 48WCAU Y 1 Surg GC- Ben CB GCI 41- F 75 WCAU N — Surg GC- Ben CN GCI 42- 0M 45 WCAU N — Surg GC-NPS CN Asterand 43- 0 F 46 CAU NU Surg 2002 As-NPSCN GCI 44- 0 F 48 WCAU Y 1 Surg GC-NPS CN GCI 45- 0 F 50 WCAU N — SurgGC-NPS CN Asterand 46- 0 F 52 CAU Occ Surg 2005 As-NPS CN Asterand 47- 0F 52 CAU occ Surg 2005 As-NPS CN GCI 48- 0 M 52 WCAU N — Surg GC-NPS CNAsterand 49- 0 M 54 CAU CurU Surg 2005 As-NPS CN GCI 50- 0 F 55 WCAU N —Surg GC-NPS CN GCI 51- 0 M 55 WCAU N — Surg GC-NPS CN GCI 52- 0 F 57WCAU Y 6 Surg GC-NPS CN GCI 53- 0 F 57 WCAU Y 1 Surg GC-NPS CN GCI 54- 0M 59 WCAU Y 42  Surg GC-NPS CN GCI 55- 0 F 61 WCAU Y 3 Surg GC-NPS CNGCI 56- 0 M 61 WCAU Y — Surg GC-NPS CN GCI 57- 0 F 66 WCAU Y 4 SurgGC-NPS CN GCI 58- 0 F 66 WCAU N — Surg GC-NPS CN GCI 59- 0 M 68 WCAU N —Surg GC-NPS CN GCI 60- 0 F 69 WCAU N — Surg GC-NPS CN Asterand 61- 0 M71 CAU Occ Surg 2005 As-NPS CN Asterand 62- 0 F 74 WCAU Occ Surg 2004As-NPS CN biochain 63- M 24 (65)- Bc-NPM CN Ambion 64- F 34 (71)- Am-NPMCN biochain 65- M 58 (66)- Bc-NPM CN biochain 66- M 61 (63)- Bc-NPM CNbiochain 67- M 66 (62)- Bc-NPM CN biochain 68- F 68 (64)- Bc-NPM CNbiochain 69- M 78 (41)- Bc-NPM CN biochain 70- F&M M (67)- (26-78)&FBc-NPM (53-77).

TABLE 5_1 Key Full Name CC Colon Cancer CB Colon Benign CN Colon NormalWT Weight HT Height Aden Adenocarcinoma AI Adenocarcinoma intramucosalUA Ulcerated adenocarcinoma WP Aden Well polypoid adeocarcinoma MAMucinus adenocarcinoma TA Tubular adenocarcinoma Carc Carcinoma TS AdenTUBULOVILLOUS ADENOMA TS Aden HGD TUBULOVILLOUS ADENOMA with HIGH GRADEDYSPLASIA NC Normal Colon N-PM Normal PM N-PM P10 Normal PM (Pool 10)Diag Diagnosis Div DIVERTICULITIS Divs w/F DIV Diverticulosis with FocalDIVERTICULITIS Chr Divs Chronic diverticulosis Divs DIVERTICULOSIS WITHDIVERTICULITIS AND w/DIV . . . FOCAL ABSCESS FORMATION; NO MALIGNANCY ADw/AF Acute diverticulitis with abscess formation CU CECAL ULCERATIONDivs, PA DIVERTICULOSIS AND PERICOLIC ABSCESS Tub Aden TUBULAR ADENOMADiv/Chr Infl DIVERTICULOSIS/CHRONIC INFLAMMATION Chr Div CHRONICDIVERTICULITIS Ext Divs EXTENSIVE DIVERTICULOSIS Divs, DIVERTICULOSISAND CHRONIC Chr Div . . . DIVERTICULITIS, SEROSAL FIBROSIS AND CHRONICSEROSITIS MU w/MI Mucosal ulceration with mural inflammation UCUlcerative colitis Cec cecum Dis C DISTAL COLON Ret, Low AntRETROSIGMOID, LOW ANTERIOR Rect Col Rectosigmoidal colon Sig col Sigmodcolon Col Sig Colon Sigma RT Col RIGHT COLON Prox T Col PROXIMALTRANSVERSE COLON LT Col Left Colon Gr Grade CS Cancer Stage Ethnic BEthnic background NU Never Used Occ Occasion Cur U Current use Dr. perday Drinks per day Alc. Dur. Alcohol Duration Auto. Autopsy Surg.Surgical Exc. Y. Excision Year

Materials and Experimental Procedures Used to Obtain Expression Data RNAPreparation—

RNA was obtained from ABS (Wilmington, Del. 19801, USA,http://www.absbioreagents.com), BioChain Inst. Inc. (Hayward, Calif.94545 USA www.biochain.com), GOG for ovary samples—Pediatic CooperativeHuman Tissue Network, Gynecologic Oncology Group Tissue Bank, ChildrenHospital of Columbus (Columbus Ohio 43205 USA), Clontech (FranklinLakes, N.J. USA 07417, www.clontech.com), Ambion (Austin, Tex. 78744USA, http://www.ambion.com), Asternad (Detroit, Mich. 48202-3420, USA,www.asterand.com), and from Genomics Collaborative Inc. a Division ofSeracare (Cambridge, Mass. 02139, USA, www.genomicsinc.com).Alternatively, RNA was generated from blood cells, cell lines or tissuesamples using TRI-Reagent (Molecular Research Center), according toManufacturer's instructions. Tissue and RNA samples were obtained frompatients or from postmortem. Most total RNA samples were treated withDNaseI (Ambion).

RT PCR—Purified RNA (2-10 μg) was mixed with 300-1500 ng Random Hexamerprimers (Invitrogen) and 500 μM dNTP in a total volume of 31.2 to 156μl. The mixture was incubated for 5 mM at 65° C. and then quicklychilled on ice. Thereafter, 10-50 μl of 5× SuperscriptII first strandbuffer (Invitrogen), 4.8 to 24 μl 0.1M DTT and 80-400 units RNasin(Promega) were added, and the mixture was incubated for 10 min at 25°C., followed by further incubation at 42° C. for 2 mM. Then, 2-10 μl(400-2000 units) of SuperscriptII (Invitrogen) was added and thereaction (final volume of 50-2500) was incubated for 50 min at 42° C.and then inactivated at 70° C. for 15 min. The resulting cDNA wasdiluted 1:20 in TE buffer (10 mM Tris pH=8, 1 mM EDTA pH=8).

Real-Time RT-PCR analysis carried out as described below—cDNA (5 μl),prepared as described above, was used as a template in Real-Time PCRreactions (final volume of 20 μl) using the SYBR Green I assay (PEApplied Biosystem) with specific primers and UNG Enzyme (Eurogentech orABI or Roche). The amplification was effected as follows: 50° C. for 2min, 95° C. for 10 min, and then 40 cycles of 95° C. for 15 sec,followed by 60° C. for 1 min, following by dissociation step. Detectionwas performed by using the PE Applied Biosystem SDS 7000. The cycle inwhich the reactions achieved a threshold level of fluorescence(Ct=Threshold Cycle, described in detail below) was registered and wasused to calculate the relative transcript quantity in the RT reactions.The relative quantity was calculated using the equation Q=efficiencŷ-Ct.The efficiency of the PCR reaction was calculated from a standard curve,created by using different dilutions of several reverse transcription(RT) reactions. To minimize inherent differences in the RT reaction, theresulting relative quantities were normalized using a normalizationfactor calculated in the following way:

The expression of several housekeeping (HSKP) genes was checked on everypanel. The relative quantity (Q) of each housekeeping gene in eachsample, calculated as described above, was divided by the medianquantity of this gene in all panel samples to obtain the “relative Q relto MED”. Then, for each sample the median of the “relative Q rel to MED”of the selected housekeeping genes was calculated and served asnormalization factor of this sample for further calculations. Schematicsummary of quantitative real-time PCR analysis is presented in FIG. 1.As shown, the x-axis shows the cycle number. The CT=Threshold Cyclepoint, which is the cycle that the amplification curve crosses thefluorescence threshold that was set in the experiment. This point is acalculated cycle number in which PCR products signal is above thebackground level (passive dye ROX) and still in theGeometric/Exponential phase (as shown, once the level of fluorescencecrosses the measurement threshold, it has a geometrically increasingphase, during which measurements are most accurate, followed by a linearphase and a plateau phase; for quantitative measurements, the latter twophases do not provide accurate measurements). The y-axis shows thenormalized reporter fluorescence. It should be noted that this type ofanalysis provides relative quantification.

For each RT sample, the expression of the specific amplicon wasnormalized to the normalization factor calculated from the expression ofdifferent house keeping genes as described in section above.

These house keeping genes are different for each panel. For colonpanel—HPRT1 (GenBank Accession No. NM_(—)000194 (SEQ ID NO: 118);amplicon—HPRT1-amplicon (SEQ ID NO:181)), PBGD (GenBank Accession No.BC019323 (SEQ ID NO: 117); amplicon—PBGD-amplicon (SEQ ID NO:178)), andG6PD (GenBank Accession No. NM_(—)000402 (SEQ ID NO: 119); G6PD amplicon(SEQ ID NO: 184)). For lung panel—HPRT1 (GenBank Accession No.NM_(—)000194 (SEQ ID NO: 118); amplicon—HPRT1-amplicon (SEQ ID NO:181)),PBGD (GenBank Accession No. BC019323 (SEQ ID NO: 117);amplicon—PBGD-amplicon (SEQ ID NO:178)), SDHA (GenBank Accession No.NM_(—)004168 (SEQ ID NO: 116); amplicon—SDHA-amplicon (SEQ ID NO:175))and Ubiquitin (GenBank Accession No. BC000449 (SEQ ID NO: 115);amplicon—Ubiquitin-amplicon (SEQ ID NO: 172)). For ovary panel—SDHA(GenBank Accession No. NM_(—)004168 (SEQ ID NO: 116);amplicon—SDHA-amplicon (SEQ ID NO:175)), HPRT1 (GenBank Accession No.NM_(—)000194 (SEQ ID NO: 118); amplicon—HPRT1-amplicon (SEQ ID NO:181))and G6PD (GenBank Accession No. NM_(—)000402 (SEQ ID NO: 119); G6PDamplicon (SEQ ID NO: 184)). For normal panel—SDHA (GenBank Accession No.NM_(—)004168 (SEQ ID NO: 116); amplicon—SDHA-amplicon (SEQ ID NO:175)),Ubiquitin (GenBank Accession No. BC000449 (SEQ ID NO: 115);amplicon—Ubiquitin-amplicon (SEQ ID NO: 172)), and TATA box (GenBankAccession No. NM_(—)003194 (SEQ ID NO: 114); TATA amplicon (SEQ ID NO:169)). For blood panel—HSB1L_HUMAN (Accession No. Q9Y450) (SEQ ID NO:109), DHSA_HUMAN (SEQ ID NO: 110) (Accession No P31040), SFRS4_HUMAN(SEQ ID NO: 111) (Accession No Q08170) and SLC25A3 (Accession No Q7Z7N7)(SEQ ID NO: 112).

The sequences of the housekeeping genes measured in all the examples ofblood panel were as follows:

HSB1L_HUMAN (SEQ ID NO: 109) (Accession No. Q9Y450)T05337_seg30-34F1-Forward primer (SEQ ID NO: 152): GCTCCAGGCCATAAGGACTTCT05337_seg30-34R1 (SEQ ID NO: 153)-Reverse primer: CAGCTTCAAACTCTCCCCTGCAmplicon (SEQ ID NO: 154):GCTCCAGGCCATAAGGACTTCATTCCAAATATGATTACAGGAGCAGCCCAGGCGGATGTAGCTGTTTTAGTTGTAGATGCCAGCAGGGGAGAGTTTGAAGCT GDHSA_HUMAN (SEQ ID NO: 110) (Accession No P31040)M78124_seg45-48F1 (SEQ ID NO: 155)-Forward primer: TTCCTTGCCAGGACCTAGAGM78124_seg45-48R1-Reverse primer (SEQ ID NO: 156): CATAAACCTTTCGCCTTGACAmplicon (SEQ ID NO: 157):TTCCTTGCCAGGACCTAGAGTTTGTTCAGTTCCACCCCACAGGCATATATGGTGCTGGTTGTCTCATTACGGAAGGATGTCGTGGAGAGGGAGGCATTCTCATTAACAGTCAAGGCGAAAGGTTTATGSFRS4_HUMAN (SEQ ID NO: 111) (Accession No Q08170)HUMSRP75Aseg30-33F1 (SEQ ID NO: 158)- Forward  primer:AATTTGTCAAGTCGGTGCAGC HUMSRP75Aseg30-33R1 (SEQ ID NO: 159)- Reverse primer: TCACCCCTTCATTTTTGCGT Amplicon (SEQ ID NO: 160):AATTTGTCAAGTCGGTGCAGCTGGCAAGACCTAAAGGATTATATGCGTCAGGCAGGAGAAGTGACTTATGCAGATGCTCACAAGGGACGCAAAAATGAAGG GGTGASLC25A3 (Accession No Q7Z7N7) (SEQ ID NO: 112)SSMPCPseg24-25-29F1- Forward primer  (SEQ ID NO: 161):CCCAAAATGTATAAGGAAGAAGGC SSMPCPseg24-25-29R1- Reverse primer (SEQ ID NO: 162): TTCAAAGCAGGCGAACTTCA Amplicon (SEQ ID NO: 163):CAGCCAGGTTATGCCAACACTTTGAGGGATGCAGCTCCCAAAATGTATAAGGAAGAAGGCCTAAAAGCATTCTACAAGGGGGTTGCTCCTCTCTGGATGAGACAGATACCATACACCATGATGAAGTTCGCCTGCTTTGA

The sequences of the housekeeping genes measured in all the examples onnormal tissue samples panel were as follows:

TATA box (GenBank Accession No. NM_003194 (SEQ ID NO: 114)),TATA box Forward primer (SEQ ID NO: 167): CGGTTTGCTGCGGTAATCATTATA box Reverse primer (SEQ ID NO: 168): TTTCTTGCTGCCAGTCTGGACTATA box-amplicon (SEQ ID NO: 169):CGGTTTGCTGCGGTAATCATGAGGATAAGAGAGCCACGAACCACGGCACTGATTTTCAGTTCTGGGAAAATGGTGTGCACAGGAGCCAAGAGTGAAGAACAGTCCAGACTGGCAGCAAGAAA Ubiquitin (GenBank Accession No. BC000449(SEQ ID NO: 115)) Ubiquitn Forward primer (SEQ ID NO: 170):ATTTGGGTCGCGGTTCTTG Ubiquitin Reverse primer (SEQ ID NO: 171):TGCCTTGACATTCTCGATGGT Ubiquitin-amplicon (SEQ ID NO: 172)ATTTGGGTCGCGGTTCTTGTTTGTGGATCGCTGTGATCGTCACTTGACAATGCAGATCTTCGTGAAGACTCTGACTGGTAAGACCATCACCCTCGAGGTTGAGCCCAGTGACACCATCGAGAATGTCAAGGCASDHA (GenBank Accession No. NM_004168 (SEQ ID NO: 116))SDHA Forward primer (SEQ ID NO: 173): TGGGAACAAGAGGGCATCTGSDHA Reverse primer (SEQ ID NO: 174): CCACCACTGCATCAAATTCATGSDHA-amplicon (SEQ ID NO: 175):TGGGAACAAGAGGGCATCTGCTAAAGTTTCAGATTCCATTTCTGCTCAGTATCCAGTAGTGGATCATGAATTTGATGCAGTGGTGG

The sequences for primers and amplicons of the housekeeping genesmeasured in all the cancer examples are listed below. For colonpanel—HPRT1, PBGD and G6PD were used. For lung panel—PBGD, HPRT1,Ubiquitin and SDHA were used. For ovary panel—HPRT1, SDHA and G6PD wereused.

SDHA (GenBank Accession No. NM_004168 (SEQ ID NO: 116):SDHA Forward primer (SEQ ID NO: 173): TGGGAACAAGAGGGCATCTGSDHA Reverse primer (SEQ ID NO: 174): CCACCACTGCATCAAATTCATGSDHA-amplicon (SEQ ID NO: 175):TGGGAACAAGAGGGCATCTGCTAAAGTTTCAGATTCCATTTCTGCTCAGTATCCAGTAGTGGATCATGAATTTGATGCAGTGGTGGPBGD (GenBank Accession No. BC019323 (SEQ ID NO: 117)),PBGD Forward primer (SEQ ID NO: 176): TGAGAGTGATTCGCGTGGGPBGD Reverse primer (SEQ ID NO: 177): CCAGGGTACGAGGCTTTCAATPBGD-amplicon (SEQ ID NO: 178):TGAGAGTGATTCGCGTGGGTACCCGCAAGAGCCAGCTTGCTCGCATACAGACGGACAGTGTGGTGGCAACATTGAAAGCCTCGTACCCTGGHPRT1 (GenBank Accession No. NM_000194 (SEQ ID NO: 118)),HPRT1 Forward primer (SEQ ID NO: 179): TGACACTGGCAAAACAATGCAHPRT1 Reverse primer (SEQ ID NO: 180): GGTCCTTTTCACCAGCAAGCTHPRT1-amplicon (SEQ ID NO: 181):TGACACTGGCAAAACAATGCAGACTTTGCTTTCCTTGGTCAGGCAGTATAATCCAAAGATGGTCAAGGTCGCAAGCTTGCTGGTGAAAAGGACCG6PD (GenBank Accession No. NM_000402 (SEQ ID NO: 119))G6PD Forward primer (SEQ ID NO: 182): gaggccgtcaccaagaacatG6PD Reverse primer (SEQ ID NO: 183): ggacagccggtcagagctcG6PD-amplicon (SEQ ID NO: 184):gaggccgtcaccaagaacattcacgagtcctgcatgagccagataggctggaaccgcatcatcgtggagaagcccttcgggagggacctgcaga gctctgaccggctgtccUbiquitin (GenBank Accession No. BC000449 (SEQ ID NO: 115))Ubiquitin Forward primer (SEQ ID NO: 170): ATTTGGGTCGCGGTTCTTGUbiquitin Reverse primer (SEQ ID NO: 171): TGCCTTGACATTCTCGATGGTUbiquitin Amplicon (SEQ ID NO: 172):ATTTGGGTCGCGGTTCTTGTTTGTGGATCGCTGTGATCGTCACTTGACAATGCAGATCTTCGTGAAGACTCTGACTGGTAAGACCATCACCCTCGAGGTTGAGCCCAGTGACACCATCGAGAATGTCAAGGCA

Another methodology used to predict the expression pattern of theproteins of the invention was MED discovery engine:

MED is a platform for collection of public gene-expression data,normalization, annotation and performance of various queries. Expressiondata from the most widely used Affymetrix microarrays is downloaded fromthe Gene Expression Omnibus (GEO—www.ncbi.nlm.nih.gov/GEO). Data ismultiplicatively normalized by setting the 95 percentile to a constantvalue (normalized expression=1200), and noise is filtered by setting thelower 30% to 0. Experiments are annotated, first automatically, and thenmanually, to identify tissue and condition, and chips are groupedaccording to this annotation, and cross verification of this grouping bycomparing the overall expression pattern of the genes of each chip tothe overall average expression pattern of the genes in this group. Eachprobeset in each group is assigned an expression value which is themedian of the expressions of that probeset in all chips included in thegroup. The vector of expression of all probesets within a certain groupis the virtual chip of that group, and the collection of all suchvirtual chips is a virtual panel. The panel (or sub-panels) can bequeried to identify probesets with a required behavior (e.g. specificexpression in a sub-set of tissues, or differential expression betweendisease and healthy tissues). These probesets are linked to LEADScontigs and to RefSeqs (http://www.ncbi.nlm.nih.gov/RefSeq/) byprobe-level mapping, for further analysis.

The Affymetrix platforms that are downloaded are HG-U95A and the HG-U133family (A,B, A2.0 and PLUS 2.0). Than three virtual panels were created:U95 and U133 Plus 2.0, based on the corresponding platforms, and U133which uses the set of common probesets for HG-U133A, HG-U133A2.0 andHG-U133 PLUS 2.0+.

The results of the MED discovery engine are presented in scatter plots.The scatter plot is a compact representation of a given panel(collection of groups). The y-axis is the (normalized) expression andthe x-axis describes the groups in the panel. For each group, the medianexpression is represented by a solid marker, and the expression valuesof the different chips in the group are represented by small dashes(“-”). The groups are ordered and marked as follows—“Other” groups (e.g.benign, non-cancer diseases, etc.) with a triangle, Treated cells with asquare, Normal with a circle, Matched with a cross, and Cancer with adiamond. The number of chips in each group is also written adjacent toit's name.

Example 2 Description for Cluster AI581519

The present invention relates to VSIG1 polypeptides, novel splicevariants and diagnostics and therapeutics based thereon.

According to the present invention, Cluster AI581519 (internal ID72756422) features 10 transcripts and 2 segments of interest, the namesfor which are given in Tables 6 and 7, respectively. The selectedprotein variants are given in table 8.

TABLE 6 Transcripts of interest Transcript Name AI581519_T0 (SEQ IDNO: 1) AI581519_T1 (SEQ ID NO: 2) AI581519_T2 (SEQ ID NO: 3) AI581519_T3(SEQ ID NO: 4) AI581519_T4 (SEQ ID NO: 5) AI581519_T5 (SEQ ID NO: 6)AI581519_T6 (SEQ ID NO: 7) AI581519_T8 (SEQ ID NO: 8) AI581519_T10 (SEQID NO: 9) AI581519_T11 (SEQ ID NO: 10)

TABLE 7 Segments of interest Segment Name AI581519_N7 (SEQ ID NO: 120)AI581519_N9 (SEQ ID NO: 121)

TABLE 8 Proteins of interest Protein Name Corresponding TranscriptsAI581519_P3 (SEQ AI581519_T0 (SEQ ID NO: 1); AI581519_T1 ID NO: 11) (SEQID NO: 2); AI581519_T2 (SEQ ID NO: 3); AI581519_T3 (SEQ ID NO: 4);AI581519_T4 (SEQ ID NO: 5) AI581519_P4 (SEQ AI581519_T5 (SEQ ID NO: 6)ID NO: 12) AI581519_P5 (SEQ AI581519_T6 (SEQ ID NO: 7) ID NO: 13)AI581519_P7 (SEQ AI581519_T8 (SEQ ID NO: 8) ID NO: 14) AI581519_P9 (SEQAI581519_T10 (SEQ ID NO: 9) ID NO: 15) AI581519_P10 (SEQ AI581519_T11(SEQ ID NO: 10) ID NO: 16)

These sequences are variants of the known protein V-set andimmunoglobulin domain containing 1 (RefSeq accession identifierNP_(—)872413, synonyms: RP5-889N15.1, 1700062D20Rik, GPA34, MGC44287,dJ889N15.1), referred to herein as the previously known protein.

VSIG1 is a V-set and immunoglobulin domain containing 1 protein alsoknown as glycoprotein A34 (GPA34). This gene was originally identifiedas a transcript encoding a protein with similarity to the glycoproteinA33 (GPA33), a colon cancer antigen (Scanlan et al. Cancer Immunotherapy6:2 2006), that has 32% identity to GPA33. The authors showed that A34mRNA and protein expression is highly tissue-restricted, as it isexpressed predominantly in stomach and testis. A34 mRNA and proteinexpression was also detected in gastric cancers, esophageal carcinomas,and ovarian cancers. In their studies they did not detect A34 in lung,breast or colon carcinomas (Scanlan et al. 2006, Cancer Immunity 6: 2).

A known wild type VSIG1 nucleic acid sequence has been reported invarious patent and non-patent literature references. For example, thesequence of AI581519_P3 (SEQ ID NO:11) is disclosed in WO2004037999,referred there as glycoprotein A34 (GPA34). This PCT applicationcontains the sequence for AI581519_P3 (SEQ ID NO:11) which encodes theA34 antigen disclosed herein. The corresponding antigen A34 is indicatedto be expressed in some tested stomach cancers (29%), esophageal (63%)and to be expressed to a much lesser number extent (9%) on testedovarian cancers. The authors suggest that this antigen may be used fortherapy and may be a suitable target for antibody based cancertherapies.

WO9926972 discloses that this antigenic protein may also exhibit immunestimulating or immune suppressing activity such as for the treatment ofvarious immune deficiencies and disorders (including severe combinedimmunodeficiency (SCID)), e.g., in regulating (up or down) growth andproliferation of T and/or B lymphocytes, as well as effecting thecytolytic activity of NK cells and other cell populations.

In addition this same protein sequence, as depicted in AI581519_P3 (SEQID NO:11) herein, is mentioned in WO9960020 and US2002193567 wherein itis identified as Human secreted protein #62.

A sequence homologous to VSIG1 variant as depicted in AI581519_P4 (SEQID NO:12) herein, (containing 2 mismatches corresponding to known SNPs)is disclosed in WO2003027228 application, which purportedly discloses anextensive list of different alleged differentially expressed sequences.

Further, a sequence closely related to AI581519_P5 (SEQ ID NO:13),(containing 2 mismatches corresponding to known SNPs) is disclosed inPCT application WO2004100774, which also teaches many other purporteddifferentially expressed sequences.

Still further, a sequence closely related to AI581519_P7 (SEQ ID NO:14),(containing 1 mismatch corresponding to known SNP) is disclosed in PCTapplication WO2004048550 which similarly contains an extensive listingof alleged differentially expressed sequences.

According to the present invention, VSIG1 is predicted to be a novel B7member, based on the presence of an IgV and an IgC2 domain. A largeportion of proteins having one domain of each Ig subtype areco-stimulatory molecules. Like other known B7 members, VSIG1 is also atype I membrane protein. In the present invention several alternativespliced variants of VSIG1 were identified, as described below,containing a unique region within the ectodomain. The new variants ofVSIG1 were demonstrated in the present invention to be overexpressed inlung adenocarcinoma and ovarian cancer.

MED discovery engine described in Example 1 herein, was used to assessthe expression of VSIG1 transcripts. Expression data for Affymetrixprobe sets 234370_at representing the VSIG1 gene data is shown in FIG.2. As evident from the scatter plot, presented in FIG. 2, the expressionof VSIG1 transcripts detectable with the above probe sets was higher inlung cancer compared to normal lung samples.

As noted above, cluster AI581519 features 10 transcripts, which werelisted in Table 6 above. These transcripts encode for proteins which arevariants of protein V-set and immunoglobulin domain containing 1. Adescription of each variant protein according to the present inventionis now provided.

Variant protein AI581519_P3 (SEQ ID NO:11) according to the presentinvention has an amino acid sequence encoded by transcripts AI581519_T0(SEQ ID NO:1), AI581519_T1 (SEQ ID NO:2), AI581519_T2 (SEQ ID NO:3),AI581519_T3 (SEQ ID NO:4) and AI581519_T4 (SEQ ID NO:5).

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein AI581519_P3 (SEQ ID NO:11) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 9,(given according to their positions on the amino acid sequence, with thealternative amino acids listed).

TABLE 9 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 23 Q −> R 35 V −> 51 S −> F 90 I −> V 127 Q −> 146 S−> R 176 K −> 176 K −> E 181 D −> G 181 D −> V 189 F −> L 195 I −> T 196L −> 202 T −> 280 T −> 288 S −> G 322 P −> 344 L −> P

The variant protein has the following domains, as determined by usingInterPro. The domains are described in Table 10:

TABLE 10 InterPro domains Analysis Domain description type Positions onprotein Immunoglobulin C2 type HMMSmart 34-123, 152-218Immunoglobulin-like ProfileScan 36-120, 140-227 Immunoglobulin-likeHMMPfam 36-118, 154-213 Immunoglobulin subtype HMMSmart 28-137, 146-229Myelin P0 protein FPrintScan  35-59, 110-139 Immunoglobulin V-setHMMPfam 21-137 Immunoglobulin V-type HMMSmart 38-118

Variant protein AI581519_P3 (SEQ ID NO:11) is encoded by the followingtranscripts: AI581519_T0 (SEQ ID NO:1), AI581519_T1 (SEQ ID NO:2),AI581519_T2 (SEQ ID NO:3), AI581519_T3 (SEQ ID NO:4) and AI581519_T4(SEQ ID NO:5).

The coding portion of transcript AI581519_TO (SEQ ID NO:1) starts atposition 171 and ends at position 1331. The transcript also has thefollowing SNPs as listed in Table 11 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed(SEQ ID NO:11)).

TABLE 11 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 238, 438, 696, 712, 1031, 1032, 2314, 2471 T −> 275, 757C −> T 322, 452 C −> A 449, 608 C −> 549 C −> G 608 A −> 696, 776, 1010,1136 A −> T 712 T −> C 735, 754, 1201 G −> A 1679, 1800, 1867 T −> A2260 

The coding portion of transcript AI581519_T1 (SEQ ID NO:2) starts atposition 171 and ends at position 1331. The transcript also has thefollowing SNPs as listed in Table 12 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 12 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 238, 438, 696, 712, 1031, 1032, 2314, 2471 T −> 275, 757C −> T 322, 452 C −> A 449, 608 C −> 549 C −> G 608 A −> 696, 776, 1010,1136 A −> T 712 T −> C 735, 754, 1201 G −> A 1679, 1800, 1867 T −> A2260 

The coding portion of transcript AI581519_T2 (SEQ ID NO:3) starts atposition 171 and ends at position 1331. The transcript also has thefollowing SNPs as listed in Table 13 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 13 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 238, 438, 696, 712, 1031, 1032, 1782 T −> 275, 757 C −>T 322, 452 C −> A 449, 608 C −> 549 C −> G 608 A −> 696, 776, 1010, 1136A −> T 712 T −> C 735, 754, 1201 G −> A 1679 

The coding portion of transcript AI581519_T3 (SEQ ID NO:4) starts atposition 171 and ends at position 1331. The transcript also has thefollowing SNPs as listed in Table 14 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 14 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 238, 438, 696, 712, 1031, 1032, 1702 T −> 275, 757 C −>T 322, 452 C −> A 449, 608 C −> 549 C −> G 608 A −> 696, 776, 1010, 1136A −> T 712 T −> C 735, 754, 1201 G −> A 1679 

The coding portion of transcript AI581519_T4 (SEQ ID NO:5) starts atposition 171 and ends at position 1331. The transcript also has thefollowing SNPs as listed in Table 15 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 15 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 238, 438, 696, 712, 1031, 1032 T −> 275, 757 C −> T 322,452 C −> A 449, 608 C −> 549 C −> G 608 A −> 696, 776, 1010, 1136 A −> T712 T −> C 735, 754, 1201

Variant protein AI581519_P4 (SEQ ID NO:12) according to the presentinvention has an amino acid sequence encoded by transcript AI581519_T5(SEQ ID NO:6). Alignments to previously published protein sequences areshown in FIG. 3A. A brief description of the relationship of the variantprotein according to the present invention to each such aligned proteinis as follows:

2. Comparison report between AI581519_P4 (SEQ ID NO:12) and knownproteins NP_(—)872413 (SEQ ID NO: 11) and Q86XK7_HUMAN (FIG. 3A):

A. An isolated chimeric polypeptide encoding for AI581519_P4 (SEQ IDNO:12), comprising a first amino acid sequence being at least 90%homologous to MVFAFWKVFLILSCLAGQVSVVQVTIPDGFVNVTVGSNVTLICIYTTTVASREQLSIQWSFFHKKEMEPIS corresponding to amino acids 1-71 of known proteinsNP_(—)872413 and Q86XK7_HUMAN (SEQ ID NO: 11), which also corresponds toamino acids 1-71 of AI581519_P4 (SEQ ID NO:12), a second amino acidsequence being at least 70%, optionally at least 80%, preferably atleast 85%, more preferably at least 90% and most preferably at least 95,96, 97, 98 or 99% homologous to a polypeptide having the sequenceHSSCLSTEGMEEKAVGQCLKMTHVRDARGRCSWTSE (SEQ ID NO: 284) corresponding toamino acids 72-107 of AI581519_P4 (SEQ ID NO:12), and a third amino acidsequence being at least 90% homologous toIYFSQGGQAVAIGQFKDRITGSNDPGNASITISHMQPADSGIYICDVNNPPDFLGQNQGILNVSVLVKPSKPLCSVQGRPETGHTISLSCLSALGTPSPVYYWHKLEGRDIVPVKENFNPTTGILVIGNLTNFEQGYYQCTAINRLGNSSCEIDLTSSHPEVGIIVGALIGSLVGAAIIISVVCFARNKAKAKAKERNSKTIAELEPMTKINPRGESEAMPREDATQLEVTLPSSIHETGPDTIQEPDYEPKPTQEPAPEPAPGSEPMAVPDLDIELELEPETQSELEPEPEPEPESEPGVVVEPLSEDEKGVVKA corresponding to amino acids 72-387of known proteins NP_(—)872413 and Q86XK7_HUMAN (SEQ ID NO: 11), whichalso corresponds to amino acids 108-423 of AI581519_P4 (SEQ ID NO:12),wherein said first amino acid sequence, second amino acid sequence andthird amino acid sequence are contiguous and in a sequential order.

C. An isolated polypeptide encoding for an edge portion of AI581519_P4(SEQ ID NO:12), comprising an amino acid sequence being at least 70%,optionally at least about 80%, preferably at least about 85%, morepreferably at least about 90% and most preferably at least about 95, 96,97, 98 or 99% homologous to the sequenceHSSCLSTEGMEEKAVGQCLKMTHVRDARGRCSWTSE (SEQ ID NO: 284) of AI581519_P4(SEQ ID NO:12).

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein AI581519_P4 (SEQ ID NO:12) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 16,(given according to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:12)).

TABLE 16 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 23 Q −> R 35 V −> 51 S −> F 87 G −> S 95 V −> A 126I −> V 163 Q −> 182 S −> R 212 K −> 212 K −> E 217 D −> G 217 D −> V 225F −> L 231 I −> T 232 L −> 238 T −> 316 T −> 324 S −> G 358 P −> 380 L−> P

The variant protein has the following domains, as determined by usingInterPro. The domains are described in Table 17:

TABLE 17 InterPro domains Analysis Domain description type Positions onprotein Immunoglobulin C2 type HMMSmart 34-159, 188-254 Immunoglobulinsubtype HMMSmart 28-173, 182-265 Immunoglobulin-like ProfileScan 36-156,176-263 Immunoglobulin V-set HMMPfam 21-173 Immunoglobulin V-typeHMMSmart 38-154

Variant protein AI581519_P4 (SEQ ID NO:12) is encoded by the AI581519_T5(SEQ ID NO:6), for which the coding portion starts at position 171 andends at position 1439. The transcript also has the following SNPs aslisted in Table 18 (given according to their position on the nucleotidesequence, with the alternative nucleic acid listed).

TABLE 18 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 238, 546, 804, 820, 1139, 1140, 2422, 2579 T −> 275, 865C −> T 322, 560 G −> A 429, 1787, 1908, 1975 T −> C 454, 843, 862, 1309C −> A 557, 716 C −> 657 C −> G 716 A −> 804, 884, 1118, 1244 A −> T 820T −> A 2368 

Variant protein AI581519_P5 (SEQ ID NO:13) according to the presentinvention has an amino acid sequence encoded by transcript AI581519_T6(SEQ ID NO:7). Alignments to previously published protein sequences areshown in FIG. 3B. A brief description of the relationship of the variantprotein according to the present invention to each such aligned proteinis as follows:

2. Comparison report between AI581519_P5 (SEQ ID NO:13) and knownproteins NP_(—)872413 and Q86XK7_HUMAN (SEQ ID NO: 11) (FIG. 3B):

A. An isolated chimeric polypeptide encoding for AI581519_P5 (SEQ IDNO:13), comprising a first amino acid sequence being at least 90%homologous to MVFAFWKVFLILSCLAGQVSVVQVTIPDGFVNVTVGSNVTLICIYTTTVASREQLSIQWSFFHKKEMEPIS corresponding to amino acids 1-71 of known proteinsNP_(—)872413 and Q86XK7_HUMAN (SEQ ID NO: 11), which also corresponds toamino acids 1-71 of AI581519_P5 (SEQ ID NO:13), a second amino acidsequence being at least 70%, optionally at least 80%, preferably atleast 85%, more preferably at least 90% and most preferably at least 95,96, 97, 98 or 99% homologous to a polypeptide having the sequenceHSSCLSTEGMEEKAVGQCLKMTHVRDARGRCSWTSESPWEEGKWPDVEAVKG TLDGQQAELQ (SEQ IDNO: 285) corresponding to amino acids 72-133 of AI581519_P5 (SEQ IDNO:13), and a third amino acid sequence being at least 90% homologous toIYFSQGGQAVAIGQFKDRITGSNDPGNASITISHMQPADSGIYICDVNNPPDFLGQNQGILNVSVLVKPSKPLCSVQGRPETGHTISLSCLSALGTPSPVYYWHKLEGRDIVPVKENFNPTTGILVIGNLTNFEQGYYQCTAINRLGNSSCEIDLTSSHPEVGIIVGALIGSLVGAAIIISVVCFARNKAKAKAKERNSKTIAELEPMTKINPRGESEAMPREDATQLEVTLPSSIHETGPDTIQEPDYEPKPTQEPAPEPAPGSEPMAVPDLDIELELEPETQSELEPEPEPEPESEPGVVVEPLSEDEKGVVKA corresponding to amino acids 72-387of known proteins NP_(—)872413 and Q86XK7_HUMAN (SEQ ID NO: 11), whichalso corresponds to amino acids 134-449 of AI581519_P5 (SEQ ID NO:13),wherein said first amino acid sequence, second amino acid sequence andthird amino acid sequence are contiguous and in a sequential order.

C. An isolated polypeptide encoding for an edge portion of AI581519_P5(SEQ ID NO:13), comprising an amino acid sequence being at least 70%,optionally at least about 80%, preferably at least about 85%, morepreferably at least about 90% and most preferably at least about 95, 96,97, 98 or 99% homologous to the sequenceHSSCLSTEGMEEKAVGQCLKMTHVRDARGRCSWTSESPWEEGKWPDVEAVKG TLDGQQAELQ (SEQ IDNO: 285) of AI581519_P5 (SEQ ID NO:13).

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein AI581519_P5 (SEQ ID NO:13) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 19,(given according to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:13)).

TABLE 19 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 23 Q −> R 35 V −> 51 S −> F 87 G −> S 95 V −> A 152I −> V 189 Q −> 208 S −> R 238 K −> 238 K −> E 243 D −> G 243 D −> V 251F −> L 257 I −> T 258 L −> 264 T −> 342 T −> 350 S −> G 384 P −> 406 L−> P

The variant protein has the following domains, as determined by usingInterPro. The domains are described in Table 20:

TABLE 20 InterPro domains Analysis Domain description type Positions onprotein Immunoglobulin subtype HMMSmart 28-199, 208-291 ImmunoglobulinC2 type HMMSmart 34-185, 214-280 Immunoglobulin V-set HMMPfam  21-199Immunoglobulin-like ProfileScan 202-289

Variant protein AI581519_P5 (SEQ ID NO:13) is encoded by the followingtranscript AI581519_T6 (SEQ ID NO:7), for which the coding portionstarts at position 171 and ends at position 1517. The transcript alsohas the following SNPs as listed in Table 21 (given according to theirposition on the nucleotide sequence, with the alternative nucleic acidlisted).

TABLE 21 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 238, 624, 882, 898, 1217, 1218, 2500, 2657 T −> 275, 943C −> T 322, 638 G −> A 429, 1865, 1986, 2053 T −> C 454, 921, 940, 1387C −> A 635, 794 C −> 735 C −> G 794 A −> 882, 962, 1196, 1322 A −> T 898T −> A 2446 

Variant protein AI581519_P7 (SEQ ID NO:14) according to the presentinvention is encoded by transcript AI581519_T8 (SEQ ID NO:8). Alignmentsto one or more previously published protein sequences are shown in FIG.3C. A brief description of the relationship of the variant proteinaccording to the present invention to each such aligned protein is asfollows:

2. Comparison report between AI581519_P7 (SEQ ID NO:14) and knownproteins NP_(—)872413 and Q86XK7_HUMAN (SEQ ID NO: 11) (FIG. 3C):

A. An isolated chimeric polypeptide encoding for AI581519_P7 (SEQ IDNO:14), comprising a first amino acid sequence being at least 90%homologous to MVFAFWKVFLILSCLAGQVSVVQVTIPDGFVNVTVGSNVTLICIYTTTVASREQLSIQWSFFHKKEMEPISIYFSQGGQAVAIGQFKDRITGSNDP corresponding to amino acids1-96 of known proteins NP_(—)872413 and Q86XK7_HUMAN (SEQ ID NO: 11),which also corresponds to amino acids 1-96 of AI581519_P7 (SEQ IDNO:14), and a second amino acid sequence being at least 90% homologousto VKPSKPLCSVQGRPETGHTISLSCLSALGTPSPVYYWHKLEGRDIVPVKENFNPTTGILVIGNLTNFEQGYYQCTAINRLGNSSCEIDLTSSHPEVGIIVGALIGSLVGAAIIISVVCFARNKAKAKAKERNSKTIAELEPMTKINPRGESEAMPREDATQLEVTLPSSIHETGPDTIQEPDYEPKPTQEPAPEPAPGSEPMAVPDLDIELELEPETQSELEPEPEPEPESEPGVVVEPLSEDEKGVVKA corresponding to amino acids 138-387 of knownproteins NP_(—)872413 and Q86XK7_HUMAN (SEQ ID NO: 11), which alsocorresponds to amino acids 97-446 of AI581519_P7 (SEQ ID NO:14), whereinsaid first amino acid sequence and second amino acid sequence arecontiguous and in a sequential order.

C. An isolated chimeric polypeptide encoding for an edge portion ofAI581519_P7 (SEQ ID NO:14), comprising a polypeptide having a length“n”, wherein n is at least about 10 amino acids in length, optionally atleast about 20 amino acids in length, preferably at least about 30 aminoacids in length, more preferably at least about 40 amino acids in lengthand most preferably at least about 50 amino acids in length, wherein atleast two amino acids comprise PV, having a structure as follows: asequence starting from any of amino acid numbers 96−x to 96; and endingat any of amino acid numbers 97+((n−2)−x), in which x varies from 0 ton−2.

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein AI581519_P7 (SEQ ID NO:14) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 22,(given according to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:14)).

TABLE 22 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 23 Q −> R 35 V −> 51 S −> F 90 I −> V 105 S −> R 135K −> 135 K −> E 140 D −> G 140 D −> V 148 F −> L 154 I −> T 155 L −> 161T −> 239 T −> 247 S −> G 281 P −> 303 L −> P

The variant protein has the following domains, as determined by usingInterPro. The domains are described in Table 23:

TABLE 23 InterPro domains Analysis Domain description type Positions onprotein Immunoglobulin-like ProfileScan 99-186 Immunoglobulin C2 typeHMMSmart 111-177  Immunoglobulin subtype HMMSmart 28-188

Variant protein AI581519_P7 (SEQ ID NO:14) is encoded by the transcriptAI581519_T8 (SEQ ID NO:8), for which the coding portion starts atposition 171 and ends at position 1208. The transcript also has thefollowing SNPs as listed in Table 24 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 24 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 238, 438, 573, 589, 908, 909, 2191, 2348 T −> 275, 634 C−> T 322, 452 C −> A 449, 485 C −> G 485 A −> 573, 653, 887, 1013 A −> T589 T −> C 612, 631, 1078 G −> A 1556, 1677, 1744 T −> A 2137 

Variant protein AI581519_P9 (SEQ ID NO:15) according to the presentinvention has an amino acid sequence encoded by transcript AI581519_T10(SEQ ID NO:9). Alignments to one or more previously published proteinsequences are shown in FIG. 3D. A brief description of the relationshipof the variant protein according to the present invention to each suchaligned protein is as follows:

2. Comparison report between AI581519_P9 (SEQ ID NO:15) and knownproteins NP_(—)872413 and Q86XK7_HUMAN (SEQ ID NO: 11) (FIG. 3D):

A. An isolated chimeric polypeptide encoding for AI581519_P9 (SEQ IDNO:15), comprising a first amino acid sequence being at least 90%homologous to MVFAFWKVFLILSCLAGQVSVVQVTIPDGFVNVTVGSNVTLICIYTTTVASREQLSIQWSFFHKKEMEPISIYFSQGGQAVAIGQFKDRITGSNDPGNASITISHMQPADSGIYICDVNNPPDFLGQNQGILNVSVLVKPSKPLCSVQGRPETGHTISLSCLSALGTPSPVYYWHKLEGRDIVPVKENF corresponding to amino acids 1-189 of knownproteins NP_(—)872413 and Q86XK7_HUMAN (SEQ ID NO: 11), which alsocorresponds to amino acids 1-189 of AI581519_P9 (SEQ ID NO:15), and asecond amino acid sequence being at least 70%, optionally at least 80%,preferably at least 85%, more preferably at least 90% and mostpreferably at least 95, 96, 97, 98 or 99% homologous to a polypeptidehaving the sequence TNHRDFGHWKSDKF (SEQ ID NO: 286) corresponding toamino acids 190-203 of AI581519_P9 (SEQ ID NO:15), wherein said firstamino acid sequence and second amino acid sequence are contiguous and ina sequential order.

C. An isolated polypeptide encoding for an edge portion of AI581519_P9(SEQ ID NO:15), comprising an amino acid sequence being at least 70%,optionally at least about 80%, preferably at least about 85%, morepreferably at least about 90% and most preferably at least about 95, 96,97, 98 or 99% homologous to the sequence TNHRDFGHWKSDKF (SEQ ID NO: 286)of AI581519_P9 (SEQ ID NO:15).

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein AI581519_P9 (SEQ ID NO:15) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 25,(given according to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:15)).

TABLE 25 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 23 Q −> R 35 V −> 51 S −> F 90 I −> V 127 Q −> 146 S−> R 176 K −> 176 K −> E 181 D −> G 181 D −> V 189 F −> L 195 F −> 202 K−>

The variant protein has the following domains, as determined by usingInterPro. The domains are described in Table 26:

TABLE 26 InterPro domains Analysis Domain description type Positions onprotein Immunoglobulin-like HMMPfam 36-118 Myelin P0 protein FPrintScan35-59, 110-139 Immunoglobulin-like ProfileScan 36-120 Immunoglobulinsubtype HMMSmart 28-137 Immunoglobulin V-set HMMPfam 21-137Immunoglobulin V-type HMMSmart 38-118

Variant protein AI581519_P9 (SEQ ID NO:15) is encoded by the transcriptAI581519_T10 (SEQ ID NO:9), for which the coding portion starts atposition 171 and ends at position 779. The transcript also has thefollowing SNPs as listed in Table 27 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 27 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 238, 438, 696, 712, 1029, 1030, 2312, 2469 T −> 275, 755C −> T 322, 452 C −> A 449, 608 C −> 549 C −> G 608 A −> 696, 774, 1008,1134 A −> T 712 T −> C 735, 752, 1199 G −> A 1677, 1798, 1865 T −> A2258 

Variant protein AI581519_P10 (SEQ ID NO:16) according to the presentinvention has an amino acid sequence as encoded by transcriptAI581519_T11 (SEQ ID NO:10). Alignments to previously published proteinsequences are shown in FIG. 3E. A brief description of the relationshipof the variant protein according to the present invention to each suchaligned protein is as follows:

2. Comparison report between AI581519_P10 (SEQ ID NO:16) and knownproteins NP_(—)872413 and Q86XK7_HUMAN (SEQ ID NO: 11) (FIG. 3E):

A. An isolated chimeric polypeptide encoding for AI581519_P10 (SEQ IDNO:16), comprising a first amino acid sequence being at least 90%homologous to MVFAFWKVFLILSCLAGQVSVVQVTIPDGFVNVTVGSNVTLICIYTTTVASREQLSIQWSFFHKKEMEPISIYFSQGGQAVAIGQFKDRITGSNDPGNASITISHMQPADSGIYICDVNNPPDFLGQNQGILNVSVLVKPSKPLCSVQGRPETGHTISLSCLSALGTPSPVYYWHKLEGRDIVPVKENFNPTTGILVIGNLTNFEQGYYQCTAINRLGNSSCE IDLTSScorresponding to amino acids 1-229 of known proteins NP_(—)872413 andQ86XK7_HUMAN (SEQ ID NO: 11), which also corresponds to amino acids1-229 of AI581519_P10 (SEQ ID NO:16), and a second amino acid sequenceRQ (SEQ ID NO: 287) corresponding to amino acids 230-231 of AI581519_P10(SEQ ID NO:16), wherein said first amino acid sequence, second aminoacid sequence and third amino acid sequence are contiguous and in asequential order.

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein AI581519_P10 (SEQ ID NO:16) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 28,(given according to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:16)).

TABLE 28 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 23 Q −> R 35 V −> 51 S −> F 90 I −> V 127 Q −> 146 S−> R 176 K −> 176 K −> E 181 D −> G 181 D −> V 189 F −> L 195 I −> T 196L −> 202 T −>

The variant protein has the following domains, as determined by usingInterPro. The domains are described in Table 29:

TABLE 29 InterPro domains Analysis Domain description type Positions onprotein Immunoglobulin C2 type HMMSmart 34-123, 152-218 Immunoglobulinsubtype HMMSmart 28-137, 146-229 Immunoglobulin-like HMMPfam 36-118,154-113 Myelin P0 protein FPrintScan  35-59, 110-139 ImmunoglobulinV-type HMMSmart 38-118 Immunoglobulin V-set HMMPfam 21-137Immunoglobulin-like ProfileScan 36-120, 140-227

Variant protein AI581519_P10 (SEQ ID NO:16) is encoded by the transcriptAI581519_T11 (SEQ ID NO:10), for which the coding portion starts atposition 171 and ends at position 863. The transcript also has thefollowing SNPs as listed in Table 30 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 30 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 238, 438, 696, 712, 889, 890, 2172, 2329 T −> 275, 757 C−> T 322, 452 C −> A 449, 608 C −> 549 C −> G 608 A −> 696, 776, 868,994 A −> T 712 T −> C 735, 754, 1059 G −> A 1537, 1658, 1725 T −> A2118 

According to an optional embodiment of the present invention, shortsegments related to the above cluster are also provided. These segmentsare up to about 120 bp in length, and so are included in a separatedescription.

Segment cluster AI581519_N7 (SEQ ID NO:120) according to the presentinvention is supported by 6 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: AI581519_T5 (SEQ ID NO:6) and AI581519_T6 (SEQ IDNO:7). Table 31 below describes the starting and ending position of thissegment on each transcript.

TABLE 31 Segment location on transcripts Segment Segment Transcript namestarting position ending position AI581519_T5 (SEQ ID NO: 6) 384 491AI581519_T6 (SEQ ID NO: 7) 384 491

Segment cluster AI581519_N9 (SEQ ID NO:121) according to the presentinvention is supported by 1 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: AI581519_T6 (SEQ ID NO:7). Table 32 belowdescribes the starting and ending position of this segment on eachtranscript.

TABLE 32 Segment location on transcripts Segment Segment Transcript namestarting position ending position AI581519_T6 (SEQ ID NO: 7) 492 569

Expression of V-set and immunoglobulin domain containing 1 (VSIG1)AI581519 transcripts which are detectable by amplicon as depicted insequence name AI581519_seg7 (SEQ ID NO:190) in normal and cancerousOvary tissues, in normal and cancerous lung tissues and in differentnormal tissues

Expression of V-set and immunoglobulin domain containing 1 (VSIG1)transcripts detectable by or according to seg7—AI581519_seg7 (SEQ IDNO:190) amplicon and primers AI581519_seg7F1 (SEQ ID NO: 188) andAI581519_seg7R1 (SEQ ID NO: 189) was measured by real time PCR on ovarypanel, lung panel and normal panel. The samples used are detailed inTable 4, Table 3 and Table 2 above, respectively.

Ovary panel—Non-detected samples (samples no. 80, 83, 100 and 109, Table4) were assigned Ct value of 41 and were calculated accordingly. Foreach RT sample, the expression of the above amplicon was normalized tothe normalization factor calculated from the expression of several housekeeping genes as described in Example 1. The normalized quantity of eachRT sample was then divided by the median of the quantities of the normalsamples (sample numbers 52-78, Table 4 above), to obtain a value of foldup-regulation for each sample relative to median of the normal samples.

FIG. 4 is a histogram showing over expression of the above-indicatedV-set and immunoglobulin domain containing 1 (VSIG1) transcripts incancerous Ovary samples relative to the normal samples.

As is evident from FIG. 4, the expression of V-set and immunoglobulindomain containing 1 (VSIG1) transcripts detectable by the above ampliconin mucinous carcinoma and endometroid samples was significantly higherthan in the non-cancerous samples (sample numbers 52-78, Table 4 above).Notably an over-expression of at least 40 fold was found in 4 out of 12mucinous carcinoma samples and in 3 out of 10 endometroid samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. Threshold of 40 fold over expression wasfound to differentiate between mucinous carcinoma and endometroid andnormal samples with P value of 6.02e-003 and 1.54e-002, respectively aschecked by exact Fisher test. The above values demonstrate statisticalsignificance of the results.

Lung panel—For each RT sample, the expression of the above amplicon wasnormalized to the normalization factor calculated from the expression ofseveral house keeping genes as described in Example 1. The normalizedquantity of each RT sample was then divided by the median of thequantities of the normal samples (sample numbers 51-64 and 69-70 Table 3above), to obtain a value of fold up-regulation for each sample relativeto median of the normal samples.

FIG. 5 is a histogram showing over expression of the above-indicatedV-set and immunoglobulin domain containing 1 (VSIG1) transcripts incancerous Lung samples relative to the normal samples.

As is evident from FIG. 5, the expression of V-set and immunoglobulindomain containing 1 (VSIG1) transcripts detectable by the above ampliconin adenocarcinoma samples was significantly higher than in thenon-cancerous samples (sample numbers 51-64 and 69-70, Table 3 above).Notably an over-expression of at least 6 fold was found in 11 out of 23adenocarcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. The P value for the difference in theexpression levels of V-set and immunoglobulin domain containing 1(VSIG1) transcripts detectable by the above amplicon in Lungadenocarcinoma samples versus the normal tissue samples was determinedby T test as 9.36e-003. Threshold of 6 fold over expression was found todifferentiate between adenocarcinoma and normal samples with P value of8.07e-004 as checked by exact Fisher test.

The above values demonstrate statistical significance of the results.

Normal panel—For each RT sample, the expression of the above ampliconwas normalized to the normalization factor calculated from theexpression of several house keeping genes as described in Example 1. Thenormalized quantity of each RT sample was then divided by the median ofthe quantities of the ovary samples (sample numbers 31, 32, 33 and 34,Table 2 above), to obtain a value of relative expression of each samplerelative to median of the ovary samples, as shown in FIG. 6A. Thenormalized quantity of each RT sample was also divided by the median ofthe quantities of the ovary samples (sample numbers 31-34, Table 2above), to obtain a value of relative expression of each sample relativeto median of the lung samples (sample numbers 26, 28, 29 and 30, Table 2above), as shown in FIG. 6B.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: AI581519_seg7F1 (SEQ ID NO: 188) forward primer;and AI581519_seg7R1 (SEQ ID NO: 189) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: AI581519_seg7 (SEQ IDNO:190).

Forward Primer >AI581519_seg7F1 (SEQ ID NO: 188): CACAGCTCGTGCCTCAGTACTGReverse Primer >AI581519_seg7R1 (SEQ ID NO: 189): AGCTACATCTTCCCCGAGCGAmplicon >AI581519_seg7 (SEQ ID NO: 190)CACAGCTCGTGCCTCAGTACTGAGGGTATGGAGGAAAAGGCAGTCGGTCAGTGTCTAAAAATGACGCACGTAAGAGACGCTCGGGGAAGATGTAGCT

Expression of V-set and immunoglobulin domain containing 1(VSIG1)AI581519 transcripts which are detectable by amplicon as depicted insequence name AI581519_seg7-9 (SEQ ID NO: 187) in normal and cancerousovary tissues and in normal and cancerous lung tissues

Expression of V-set and immunoglobulin domain containing 1(VSIG1)transcripts detectable by or according to seg7-9—AI581519_seg7-9 (SEQ IDNO: 187) amplicon and primers AI581519_seg7-9F1 (SEQ ID NO: 185) andAI581519_seg7-9R1 (SEQ ID NO: 186) was measured by real time PCR onovary panel and lung panel. The samples used are detailed in Table 4 andTable 3 above, respectively.

Ovary panel—Non-detected sample (sample no. 40 Table 4) was assigned Ctvalue of 41 and was calculated accordingly. For each RT sample, theexpression of the above amplicon was normalized to the normalizationfactor calculated from the expression of several house keeping genes asdescribed in Example 1. The normalized quantity of each RT sample wasthen divided by the median of the quantities of the normal samples(sample numbers 52-78, Table 4 above), to obtain a value of foldup-regulation for each sample relative to median of the normal samples.

FIG. 7 is a histogram showing over expression of the above-indicatedV-set and immunoglobulin domain containing 1(VSIG1) transcripts incancerous Ovary samples relative to the normal samples.

As is evident from FIG. 7, the expression of V-set and immunoglobulindomain containing 1(VSIG1) transcripts detectable by the above ampliconin mucinous carcinoma samples was significantly higher than in thenon-cancerous samples (sample numbers 52-78, Table 4 above) and washigher in a few adenocarcinoma samples than in the non-canceroussamples. Notably an over-expression of at least 6 fold was found in 6out of 9 mucinous carcinoma samples and in 9 out of 37 endometroidsamples.

Statistical analysis was applied to verify the significance of theseresults, as described below. Threshold of 6 fold over expression wasfound to differentiate between mucinous carcinoma and normal sampleswith P value of 2.75e-004 as checked by exact Fisher test. Threshold of6 fold over expression was found to differentiate between adenocarcinomaand normal samples with P value of 2.44e-002 as checked by exact Fishertest.

The above values demonstrate statistical significance of the results.

Lung panel—For each RT sample, the expression of the above amplicon wasnormalized to the normalization factor calculated from the expression ofseveral house keeping genes as described in Example 1. The normalizedquantity of each RT sample was then divided by the median of thequantities of the normal samples (sample numbers 51-64 and 69-70, Table3 above), to obtain a value of fold up-regulation for each samplerelative to median of the normal samples.

FIG. 8 is a histogram showing over expression of the above-indicatedV-set and immunoglobulin domain containing 1(VSIG1) transcripts incancerous Lung samples relative to the normal samples.

As is evident from FIG. 8, the expression of V-set and immunoglobulindomain containing 1(VSIG1) transcripts detectable by the above ampliconin adenocarcinoma samples was significantly higher than in thenon-cancerous samples (sample numbers 51-64 and 69-70, Table 3 above)and was higher in a few non-small cell carcinoma samples than in thenon-cancerous samples. Notably an over-expression of at least 17 foldwas found in 8 out of 18 adenocarcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. The P value for the difference in theexpression levels of V-set and immunoglobulin domain containing 1(VSIG1)transcripts detectable by the above amplicon in Lung adenocarcinomasamples versus the normal tissue samples was determined by T test as1.17e-002.

Threshold of 17 fold over expression was found to differentiate betweenadenocarcinoma and normal samples with P value of 2.41e-003 as checkedby exact Fisher test.

The above values demonstrate statistical significance of the results.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: AI581519_seg7-9F1 (SEQ ID NO: 185) forward primer;and AI581519_seg7-9R1 (SEQ ID NO: 186) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: AI581519_seg7-9 (SEQID NO: 187).

Forward Primer >AI581519_seg7-9F1 (SEQ ID NO: 185):AATGACGCACGTAAGAGACGCReverse Primer >AI581519_seg7-9R1 (SEQ ID NO: 186): GAGTGCCCTTCACAGCCTCAAmplicon >AI581519_seg7-9 (SEQ ID NO: 187)AATGACGCACGTAAGAGACGCTCGGGGAAGATGTAGCTGGACCTCTGAGTCTCCTTGGGAGGAGGGGAAGTGGCCAGATGTTGAGGCTGTGAAGGGCACTC

Expression of V-set and immunoglobulin domain containing 1(VSIG1)AI581519 transcripts which are detectable by amplicon as depicted insequence name AI581519seg7-9 (SEQ ID NO: 196) in the blood-specificpanel.

Expression of VSIG1 transcripts detectable by or according toseg7-9-AI581519seg7-9F3R3 (SEQ ID NO:196) amplicon and primersAI581519seg7-9F3 (SEQ ID NO:194) and AI581519seg7-9R3 (SEQ ID NO:195)was measured by real time PCR on blood panel. The samples used aredetailed in Table 1 above. Non-detected samples (samples no. 28, 33, 83,85, 90 and 63, Table 1) were assigned Ct value of 41 and were calculatedaccordingly. The samples used are detailed in Table 1 above. For each RTsample, the expression of the above amplicon was normalized to thenormalization factor calculated from the expression of several housekeeping genes as described in Example 1. The normalized quantity of eachRT sample was then divided by the median of the quantities of the normalsamples (sample numbers 64, 69-72 and 74-76, Table 1 above), to obtain avalue of relative expression of each sample relative to median of thenormal samples.

The results of this analysis are depicted in the histogram in FIG. 9.Expression of the above-indicated VSIG1 transcript is high in CD8, CD4untreated and CD4+CD25− samples but even higher in normal smallintestine and normal stomach samples.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: seg7-9F3 forward primer (SEQ ID NO:194); andseg7-9R3 reverse primer (SEQ ID NO:195).

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: seg7-9F3R3 (SEQ IDNO:196).

Forward Primer >AI581519seg7-9F3 (SEQ ID NO: 194)ATGACGCACGTAAGAGACGCTCG Reverse Primer >AI581519seg7-9R3(SEQ ID NO: 195) GGAGTTCAGCCTGCTGTCCATCAAG Amplicon >AI581519seg7-9F3R3(SEQ ID NO: 196) ATGACGCACGTAAGAGACGCTCGGGGAAGATGTAGCTGGACCTCTGAGTCTCCTTGGGAGGAGGGGAAGTGGCCAGATGTTGAGGCTGTGAAGGGCACTCTTGATGGACAGCAGGCTGAACTCC

Expression of V-set and immunoglobulin domain containing 1(VSIG1)AI581519 transcripts which are detectable by amplicon as depicted insequence name AI581519_junc7-11F2R2 (SEQ ID NO:193) in normal andcancerous lung tissues, normal and cancerous ovary tissues, differentnormal tissues and blood-specific panel.

Expression of VSIG1 transcripts detectable by or according tojunc7-11F2R2-AI581519_junc7-11F2R2 (SEQ ID NO:193) amplicon and primersAI581519_junc7-11F2 (SEQ ID NO:191) and AI581519_junc7-11R2 (SEQ IDNO:192) was measured by real time PCR on lung panel, ovary panel, normalpanel and blood panel. The samples used are detailed in Table 3, Table4, Table 2 and Table 1 above, respectively. For each RT sample, theexpression of the above amplicon was normalized to the normalizationfactor calculated from the expression of several house keeping genes asdescribed in Example 1.

For lung panel—Non-detected sample (sample no. 49, Table 3) was assignedCt value of 41 and was calculated accordingly. The normalized quantityof each RT sample was then divided by the median of the quantities ofthe normal samples (samples numbers 51-64, 69 and 70, Table 3 above), toobtain a value of fold up-regulation for each sample relative to medianof the normal samples.

FIG. 10 is a histogram showing over expression of the above-indicatedVSIG1 transcripts in cancerous Lung samples relative to the normalsamples.

As is evident from FIG. 10, the expression of VSIG1 transcriptsdetectable by the above amplicon in adenocarcinoma samples wassignificantly higher than in the non-cancerous samples (sample numbers51-64, 69 and 70, Table 3 above). Notably an over-expression of at least16 fold was found in 10 out of 23 adenocarcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. The P value for the difference in theexpression levels of VSIG1 transcripts detectable by the above ampliconin lung adenocarcinoma samples versus the normal tissue samples wasdetermined by T test as 5.13e-003.

Threshold of 16 fold over expression was found to differentiate betweenadenocarcinoma and normal samples with P value of 1.80e-003 as checkedby exact Fisher test.

The above values demonstrate statistical significance of the results.

For ovary panel—Non-detected samples (samples no. 16, 23, 57, 60-62, 67,68, 71-74, 77 and 78, Table 4) were assigned Ct value of 41 and werecalculated accordingly. The normalized quantity of each RT sample wasthen divided by the median of the quantities of the normal samples(sample numbers 52, 53, 55 and 57-67, Table 4 above), to obtain a valueof fold up-regulation for each sample relative to median of the normalsamples.

FIG. 11 is a histogram showing over expression of the above-indicatedVSIG1 transcripts in cancerous Ovary samples relative to the normalsamples.

As is evident from FIG. 11, the expression of VSIG1 transcriptsdetectable by the above amplicon in mucinous carcinoma samples wassignificantly higher than in the non-cancerous samples (sample numbers52, 53, 55 and 57-67, Table 4 above) and was higher in a fewadenocarcinoma samples than in the non-cancerous samples. Notably anover-expression of at least 25 fold was found in 6 out of 9 mucinouscarcinoma samples and in 10 out of 37 adenocarcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. Threshold of 25 fold over expression wasfound to differentiate between mucinous carcinoma samples andadenocarcinoma and normal samples with P value of 3.95e-004 and1.88e-002, respectively, as checked by exact Fisher test. The abovevalues demonstrate statistical significance of the results.

For normal panel—Non-detected samples (samples no. 11-20, 28, 30, 32-34,36, 38-40, 49 and 56, Table 2) were assigned Ct value of 41 and werecalculated accordingly. The normalized quantity of each RT sample wasthen divided by the median of the quantities of the lung samples (samplenumbers 26, and 28-30, Table 2 above), to obtain a value of relativeexpression of each sample relative to median of the lung samples, asshown in FIG. 12A. The normalized quantity of each RT sample was thendivided by the median of the quantities of the ovary samples (samplenumbers 31-34, Table 2 above), to obtain a value of relative expressionof each sample relative to median of the ovary samples, as shown in FIG.12B.

For blood panel—Non-detected samples (samples no. 6, 15-19, 22-24, 27,29, 40, 41, 46-50, 52-58, 60-64, 71 and 76, Table 1) were assigned Ctvalue of 41 and were calculated accordingly. The normalized quantity ofeach RT sample was then divided by the median of the quantities of thekidney normal samples (sample numbers 65-67, Table 1 above), to obtain avalue of relative expression of each sample relative to median of thenormal samples.

The results of this analysis are depicted in the histogram in FIG. 13.Expression of the above-indicated VSIG1 transcript is very high in CD8,CD4 untreated and CD4+CD25− samples, is high in several lymphomas butalso very high in normal small intestine and normal stomach samples.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: AI581519 junc7-11F2 (SEQ ID NO:191) forwardprimer; and AI581519 junc7-11R2 (SEQ ID NO:192) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: AI581519 junc7-11F2R2(SEQ ID NO:193).

Forward Primer >AI581519_junc7-11F2 (SEQ ID NO: 191)GAAGATGTAGCTGGACCTCTGAGATTTA Reverse Primer >AI581519_junc7-11R2(SEQ ID NO: 192) GTTGGACCCTGTAATTCGATCTTTAmplicon >AI581519_junc7-11F2R2 (SEQ ID NO: 193)GAAGATGTAGCTGGACCTCTGAGATTTACTTTTCTCAAGGTGGACAAGCTGTAGCCATCGGGCAATTTAAAGATCGAATTACAGGGTCCAAC

Example 3 Description for Cluster AA424839

The present invention relates to ILDR1 polypeptides, novel splicevariants and diagnostics and therapeutics based thereon.

According to the present invention, Cluster AA424839 (internal ID71418261) features 4 transcripts and 1 segment of interest, the namesfor which are given in Tables 33 and 34, respectively. The selectedprotein variants are given in table 35.

TABLE 33 Transcripts of interest Transcript Name AA424839_T0 (SEQ ID NO:17) AA424839_T2 (SEQ ID NO: 18) AA424839_T4 (SEQ ID NO: 19)AA424839_1_T7 (SEQ ID NO: 20)

TABLE 34 Segments of interest Segment Name AA424839_N18 (SEQ ID NO: 122)

TABLE 35 Proteins of interest Protein Name Corresponding TranscriptsAA424839_P3 (SEQ ID NO: 22) AA424839_T0 (SEQ ID NO: 17) AA424839_P5 (SEQID NO: 21) AA424839_T2 (SEQ ID NO: 18) AA424839_P7 (SEQ ID NO: 23)AA424839_T4 (SEQ ID NO: 19) AA424839_1_P11 (SEQ ID NO: AA424839_1_T7(SEQ ID NO: 20) 24)

These sequences are variants of the known protein immunoglobulin-likedomain containing receptor 1 (RefSeq accession identifier NP_(—)787120(SEQ ID NO: 21), also known as ILDR1alpha, ILDR1beta, MGC50831),referred to herein as the previously known protein.

ILDR1, denoted immunoglobulin-like domain containing receptor 1 (SEQ IDNO:21), was described by Hauge et al. (2004) BBRC 323: 970-978, thatdemonstrated differential expression of the transcripts encoding thisprotein in indolent follicular lymphoma (FL) and matched transformeddiffuse large B cell lymphoma (DLBCL). The gene was identified using acDNA subtraction strategy on patient-matched biopsies of FL and DLBCL.The protein was shown to contain a signal peptide and transmembranedomain, and an Ig domain in the extracellular portion, and it was foundto be membrane bound protein, having 31% identity tolipolysis-stimulated remnant receptor (LSR).

According to the present invention, ILDR1 protein and ILDR1 splicevariants were predicted to be novel B7/CD28 members. According to thepresent invention, ILDR1 and ILDR1 splice variants were demonstrated tobe overexpressed in ovarian cancer.

MED discovery engine described in Example 1 herein, was used to assessthe expression of ILDR1 transcripts. Expression data for Affymetrixprobe set 235583_at representing the ILDR1 gene data is shown in FIGS.14 and 15. As evident from the scatter plot, presented in FIG. 14, theexpression of ILDR1 transcripts detectable with the above probe sets washigher in ovarian cancer compared to normal ovary samples. As evidentfrom the scatter plot, presented in FIG. 15, the expression of ILDR1transcripts detectable with the above probe sets was higher in coloncancer compared to normal colon samples.

As noted above, cluster AA424839 features 4 transcripts, which werelisted in Table 33 above. These transcripts encode for proteins whichare variants of protein immunoglobulin-like domain containing receptor 1(SEQ ID NO:21). A description of each protein according to the presentinvention is now provided.

Variant protein AA424839_P3 (SEQ ID NO:22) according to the presentinvention has an amino acid sequence as encoded by transcriptAA424839_TO (SEQ ID NO:17). Alignments to previously published proteinsequences are shown in FIGS. 16A. A brief description of therelationship of the variant protein according to the present inventionto each such aligned protein is as follows:

1. Comparison report between AA424839_P3 (SEQ ID NO:22) and knownproteins Q86SU0_HUMAN (SEQ ID NO: 21) and NP_(—)787120 (SEQ ID NO: 21)(FIG. 16A):

A. An isolated chimeric polypeptide encoding for AA424839_P3 (SEQ IDNO:22), comprising a first amino acid sequence being at least 90%homologous to MAWPKLPAPWLLLCTWLPAGCLSLLVTVQHTERYVTLFASIILKCDYTTSAQLQDVVVTWRFKSFCKDPIFDYYSASYQAALSLGQDPSNDCNDNQREVRIVAQRRGQNEPVLGVDYRQRKITIQNRADLVINEVMWWDHGVYYCTIEAPGDTSGDPDKEVKLIVLHWLTVIFIILGALLLLLLIGVCWCQCCPQYCCCYIRCPCCPAHCCCPEE corresponding toamino acids 1-215 of known proteins Q86SU0_HUMAN (SEQ ID NO: 21) andNP_(—)787120 (SEQ ID NO: 21), which also corresponds to amino acids1-215 of AA424839_P3 (SEQ ID NO:22), a second amino acid sequence beingat least 70%, optionally at least 80%, preferably at least 85%, morepreferably at least 90% and most preferably at least 95, 96, 97, 98 or99% homologous to a polypeptide having the sequenceALARHRYMKQAQALGPQMMGKPLYWGADRSSQVSSYPMHPLLQR (SEQ ID NO: 288)corresponding to amino acids 216-259 of AA424839_P3 (SEQ ID NO:22), anda third amino acid sequence being at least 90% homologous toDLSLPSSLPQMPMTQTTNQPPIANGVLEYLEKELRNLNLAQPLPPDLKGRFGHPCSMLSSLGSEVVERRIIHLPPLIRDLSSSRRTSDSLHQQWLTPIPSRPWDLREGRSHHHYPDFHQELQDRGPKSWALERRELDPSWSGRHRSSRLNGSPIHWSDRDSLSDVPSSSEARWRPSHPPFRSRCQERPRRPSPRESTQRHGRRRRHRSYSPPLPSGLSSWSSEEDKERQPQSWRAHRRGSHSPHWPEEKPPSYRSLDITPGKNSRKKGSVERRSEK DSSHSGRSVVIcorresponding to amino acids 216-502 of known proteins Q86SU0_HUMAN (SEQID NO: 21) and NP_(—)787120 (SEQ ID NO: 21), which also corresponds toamino acids 260-546 of AA424839_P3 (SEQ ID NO:22), wherein said firstamino acid sequence, second amino acid sequence and third amino acidsequence are contiguous and in a sequential order.

B. An isolated polypeptide encoding for an edge portion of AA424839_P3(SEQ ID NO:22), comprising an amino acid sequence being at least 70%,optionally at least about 80%, preferably at least about 85%, morepreferably at least about 90% and most preferably at least about 95, 96,97, 98 or 99% homologous to the sequenceALARHRYMKQAQALGPQMMGKPLYWGADRSSQVSSYPMHPLLQR (SEQ ID NO: 288) ofAA424839_P3 (SEQ ID NO:22).

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein AA424839_P3 (SEQ ID NO:22) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 36,(given according to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:22)).

TABLE 36 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 165 V −> F 165 V −> L 264 P −> R 388 W −> L 388 W −>S 436 H −> N 500 H −> L 500 H −> P 516 D −> Y

The variant protein has the following domains, as determined by usingInterPro. The domains are described in Table 37.

TABLE 37 InterPro domains Analysis Domain description type Positions onprotein Phospholipase A2 ScanRegExp 205-212 Immunoglobulin subtypeHMMSmart  30-166

Variant protein AA424839_P3 (SEQ ID NO:22) is encoded by the transcriptAA424839_T0 (SEQ ID NO:17), for which the coding portion starts atposition 204 and ends at position 1841. The transcript also has thefollowing SNPs as listed in Table 38 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 38 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence G −> T 696, 1366, 1748, 1749 G −> C 696, 995, 1366 C −> G 994,1529, 2305 A −> T 1442, 1702 A −> C 1442, 1702 G −> A 1502 C −> A 1509,1529

Protein AA424839_P5 (SEQ ID NO:21) according to the present inventionhas an amino acid sequence as encoded by transcript AA424839_T2 (SEQ IDNO:18).

The localization of the protein was determined according to results froma number of different software programs and analyses, including analysesfrom SignalP and other specialized programs. The protein is believed tobe located as follows with regard to the cell: membrane.

Protein AA424839_P5 (SEQ ID NO:21) also has the following non-silentSNPs (Single Nucleotide Polymorphisms) as listed in Table 39, (givenaccording to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:21)).

TABLE 39 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 165 V −> F 165 V −> L 220 P −> R 344 W −> L 344 W −>S 392 H −> N 456 H −> L 456 H −> P 472 D −> Y

The protein has the following domains, as determined by using InterPro.The domains are described in Table 40:

TABLE 40 InterPro domains Analysis Domain description type Positions onprotein Phospholipase A2 ScanRegExp 205-212 Immunoglobulin subtypeHMMSmart  30-166

Protein AA424839_P5 (SEQ ID NO:21) is encoded by the transcriptAA424839_T2 (SEQ ID NO:18), for which the coding portion starts atposition 204 and ends at position 1709. The transcript also has thefollowing SNPs as listed in Table 41 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 41 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence G −> T 696, 1234, 1616, 1617 G −> C 696, 863, 1234 C −> G 862,1397, 2173 A −> T 1310, 1570 A −> C 1310, 1570 G −> A 1370 C −> A 1377,1397

The genomic structure of protein AA424839_P5 (SEQ ID NO:21) (number ofexons relevant to the extra-cellular region of the protein, the lengthof these exons, the frame of the codon in which the introns are insertedand the location of the protein features and domains in the genestructure) is characteristic to the ligands of the B7/co-stimulatoryprotein family, as given in table 42

TABLE 42 genomic structure and protein features Exon Exon Amino- numberLength Acids Protein feature on exon 1 58  1-19 Signal Peptide 2 17120-76 Ig Domain 3 150  77-126 Ig Domain 4 120 127-166 IgDomain/Trans-membrane region 5 147 167-215 Trans-membrane region 6 821216-489 7 39 490-502

Variant protein AA424839_P7 (SEQ ID NO:23) according to the presentinvention has an amino acid sequence as encoded by transcriptAA424839_T4 (SEQ ID NO:19). Alignments to one or more previouslypublished protein sequences are shown in FIG. 16B. A brief descriptionof the relationship of the variant protein according to the presentinvention to each such aligned protein is as follows:

1. Comparison report between AA424839_P7 (SEQ ID NO:23) and knownproteins Q86SU0_HUMAN and NP_(—)787120 (SEQ ID NO: 21) (FIG. 16B):

A. An isolated chimeric polypeptide encoding for AA424839_P7 (SEQ IDNO:23), comprising a first amino acid sequence being at least 90%homologous to MAWPKLPAPWLLLCTWLPAGCLSLLVTVQHTERYVTLFASIILKCDYTTSAQLQDVVVTWRFKSFCKDPIFDYYSASYQAALSLGQDPSNDCNDNQREVRIVAQRRGQNEPVLGVDYRQRKITIQN corresponding to amino acids 1-126 of known proteinsQ86SU0_HUMAN and NP_(—)787120 (SEQ ID NO: 21), which also corresponds toamino acids 1-126 of AA424839_P7 (SEQ ID NO:23), a second amino acidsequence being at least 70%, optionally at least 80%, preferably atleast 85%, more preferably at least 90% and most preferably at least 95,96, 97, 98 or 99%, homologous to a polypeptide having the sequencePLARHRYMKQAQALGPQMMGKPLYWGADRSSQVSSYPMHPLLQR corresponding to aminoacids 127-170 of AA424839_P7 (SEQ ID NO:23), and a third amino acidsequence being at least 90% homologous toDLSLPSSLPQMPMTQTTNQPPIANGVLEYLEKELRNLNLAQPLPPDLKGRFGHPCSMLSSLGSEVVERRIIHLPPLIRDLSSSRRTSDSLHQQWLTPIPSRPWDLREGRSHHHYPDFHQELQDRGPKSWALERRELDPSWSGRHRSSRLNGSPIHWSDRDSLSDVPSSSEARWRPSHPPFRSRCQERPRRPSPRESTQRHGRRRRHRSYSPPLPSGLSSWSSEEDKERQPQSWRAHRRGSHSPHWPEEKPPSYRSLDITPGKNSRKKGSVERRSEK DSSHSGRSVVIcorresponding to amino acids 216-502 of known proteins Q86SU0_HUMAN andNP_(—)787120 (SEQ ID NO: 21), which also corresponds to amino acids171-457 of AA424839_P7 (SEQ ID NO:23), wherein said first amino acidsequence, second amino acid sequence and third amino acid sequence arecontiguous and in a sequential order.

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: secreted.

Variant protein AA424839_P7 (SEQ ID NO:23) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 43,(given according to their positions on the amino acid sequence, with thealternative amino acids listed).

TABLE 43 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 175 P −> R 299 W −> L 299 W −> S 347 H −> N 411 H −>L 411 H −> P 427 D −> Y

Variant protein AA424839_P7 (SEQ ID NO:23) is encoded by the transcriptAA424839_T4 (SEQ ID NO:19), for which the coding portion starts atposition 204 and ends at position 1574. The transcript also has thefollowing SNPs as listed in Table 44 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 44 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> G 727, 1262, 2038 G −> C  728, 1099 G −> T 1099, 1481,1482 A −> T 1175, 1435 A −> C 1175, 1435 G −> A 1235 C −> A 1242, 1262

Variant protein AA424839_(—)1_P11 (SEQ ID NO:24) according to thepresent invention has an amino acid sequence as encoded by transcriptAA424839_(—)1_T7 (SEQ ID NO:20). Alignments to one or more previouslypublished protein sequences are given in FIG. 16C.

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein AA424839_(—)1_P11 (SEQ ID NO:24) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 45,(given according to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:24)).

TABLE 45 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 108 V −> F 108 V −> L 207 P −> R 331 W −> L 331 W −>S 379 H −> N 443 H −> L 443 H −> P 459 D −> Y

The variant protein has the following domains, as determined by usingInterPro. The domains are described in Table 46:

TABLE 46 InterPro domains Analysis Domain description type Positions onprotein Phospholipase A2 ScanRegExp 148-155 Immunoglobulin subtypeHMMSmart  30-109

Variant protein AA424839_(—)1_P11 (SEQ ID NO:24) is encoded by thetranscript AA424839_(—)1_T7 (SEQ ID NO:20), for which the coding portionstarts at position 204 and ends at position 1670. The transcript alsohas the following SNPs as listed in Table 47 (given according to theirposition on the nucleotide sequence, with the alternative nucleic acidlisted).

TABLE 47 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence G −> T 525, 1195, 1577, 1578 G −> C 525, 824, 1195 C −> G 823,1358, 2134 A −> T 1271, 1531 A −> C 1271, 1531 G −> A 1331 C −> A 1338,1358

Segment cluster AA424839_N18 (SEQ ID NO:122) according to the presentinvention is supported by 10 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: AA424839_TO (SEQ ID NO:17), AA424839_T2 (SEQ IDNO:18), AA424839_T4 (SEQ ID NO:19) and AA424839_(—)1_T7 (SEQ ID NO:20).Table 48 below describes the starting and ending position of thissegment on each transcript.

TABLE 48 Segment location on transcripts Segment Segment Transcript namestarting position ending position AA424839_T0 (SEQ ID NO: 17) 1173 1802AA424839_T2 (SEQ ID NO: 18) 1041 1670 AA424839_T4 (SEQ ID NO: 19) 9061535 AA424839_1_T7 (SEQ ID NO: 20) 1002 1631

Expression of immunoglobulin-like domain containing receptor 1 (ILDR1)AA424839 transcripts which are detectable by amplicon as depicted insequence name AA424839_seg18wt (SEQ ID NO:199) in normal and cancerousOvary tissues and in different normal tissues

Expression of immunoglobulin-like domain containing receptor 1 (ILDR1)transcripts detectable by or according to seg18wt—AA424839_seg18wt (SEQID NO:199) amplicon and primers AA424839_seg18wtF1 (SEQ ID NO:197) andAA424839_seg18wtR1 (SEQ ID NO:198) was measured by real time PCR onovary panel and normal panel. The samples used are detailed in Table 4and Table 2 above, respectively. For each RT sample, the expression ofthe above amplicon was normalized to the normalization factor calculatedfrom the expression of several house keeping genes as described inExample 1.

Ovary panel—The normalized quantity of each RT sample was then dividedby the median of the quantities of the normal samples (sample numbers52-78, Table 4 above), to obtain a value of fold up-regulation for eachsample relative to median of the normal samples.

FIG. 17 is a histogram showing over expression of the above-indicatedimmunoglobulin-like domain containing receptor 1 (ILDR1) transcripts incancerous Ovary samples relative to the normal samples.

As is evident from FIG. 17, the expression of immunoglobulin-like domaincontaining receptor 1 (ILDR1) transcripts detectable by the aboveamplicon in serous carcinoma, mucinous carcinoma and adenocarcinomasamples was significantly higher than in the non-cancerous samples(sample numbers 52-78, Table 4 above). Notably an over-expression of atleast 25 fold was found in 36 out of 37 adenocarcinoma samples,specifically in 17 out of 18 serous carcinoma samples, in 9 out of 9mucinous carcinoma samples and in 10 out of 10 endometroid samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. The P value for the difference in theexpression levels of immunoglobulin-like domain containing receptor 1(ILDR1) transcripts detectable by the above amplicon in Ovaryadenocarcinoma samples, serous carcinoma samples mucinous carcinoma andendometriod versus the normal tissue samples was determined by T test as3.85e-010, 6.21e-005, 1.10e-003 and 2.94e-004 respectively.

Threshold of 25 fold over expression was found to differentiate betweenadenocarcinoma, serous carcinoma, mucinous carcinoma, endometriod andnormal samples with P value of 3.31e-017, 1.63e-011, 1.06e-008 and2.87e-009, respectively, as checked by exact Fisher test.

The above values demonstrate statistical significance of the results.

Normal panel—The normalized quantity of each RT sample was then dividedby the median of the quantities of the ovary samples (sample numbers 31,32, 33 and 34, Table 2 above), to obtain a value of relative expressionof each sample relative to median of the ovary samples, as shown in FIG.18.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: AA424839_seg18wtF1 (SEQ ID NO:197) forward primer;and AA424839_seg18wtR1 (SEQ ID NO:198) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: AA424839_seg18wt (SEQID NO:199).

Forward Primer >AA424839_seg18wtF1 (SEQ ID NO: 197) AGCCACCACCATTACCCTGAReverse Primer >AA424839_seg18wtR1 (SEQ ID NO: 198) TGCCTTCCACTCCACGATGAmplicon >AA424839_seg18wt (SEQ ID NO: 199)AGCCACCACCATTACCCTGATTTCCACCAGGAGCTCCAGGACCGGGGGCCAAAGTCTTGGGCATTGGAAAGAAGGGAGTTGGACCCATCGTGGA GTGGAAGGCA

Expression of immunoglobulin-like domain containing receptor 1 (ILDR1)AA424839 transcripts which are detectable by amplicon as depicted insequence name AA424839_seg14-16 (SEQ ID NO: 202) in normal and cancerousovary tissues or different normal tissues

Expression of ILDR1 transcripts detectable by or according toseg14-16-AA424839_seg14-16 (SEQ ID NO: 202) amplicon and primersAA424839_seg14-16E1 (SEQ ID NO:200) and AA424839_seg14-16R1 (SEQ IDNO:201) was measured by real time PCR on ovary panel or normal panel.The samples used are detailed in Table 4 and in table 2 above,respectively. For each RT sample, the expression of the above ampliconwas normalized to the normalization factor calculated from theexpression of several house keeping genes as described in Example 1.

For ovary panel—the normalized quantity of each RT sample was thendivided by the median of the quantities of the normal samples (samplenumbers 52-55, 58, 59, 63-69 and 71-78, Table 4 above), to obtain avalue of fold up-regulation for each sample relative to median of thenormal samples.

FIG. 19 is a histogram showing over expression of the above-indicatedILDR1 transcripts in cancerous Ovary samples relative to the normalsamples.

As is evident from FIG. 19, the expression of ILDR1 transcriptsdetectable by the above amplicon in serous carcinoma, mucinous carcinomaand adenocarcinoma samples was significantly higher than in thenon-cancerous samples (sample numbers 52-55, 58, 59, 63-69 and 71-78,Table 4 above). Notably an over-expression of at least 14 fold was foundin 33 out of 37 adenocarcinoma samples: 14 out of 18 serous carcinomasamples, in 9 out of 9 mucinous carcinoma samples and in 10 out of 10adenocarcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below.

The P value for the difference in the expression levels of ILDR1transcripts detectable by the above amplicon in ovary adenocarcinomasamples, ovary serous carcinoma samples, ovary mucinous carcinomasamples and ovary endometroid samples versus the normal tissue sampleswas determined by T test as 1.00e-008, 4.79e-004, 4.97e-004 and6.93e-005, respectively.

Threshold of 14 fold over expression was found to differentiate betweenadenocarcinoma, serous carcinoma, mucinous carcinoma, endometriod andnormal samples with P value of 3.78e-012, 2.03e-007, 6.99e-008 and2.25e-008, respectively, as checked by exact Fisher test.

The above values demonstrate statistical significance of the results.

For normal panel—The normalized quantity of each RT sample was thendivided by the median of the quantities of the ovary samples (samplenumbers 31-34, Table 2 above), to obtain a value of relative expressionof each sample relative to median of the ovary samples.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: AA424839_seg14-16F1 (SEQ ID NO:200) forwardprimer; and AA424839_seg14-16R1 (SEQ ID NO:201) reverse primer.

The results demonstrating the expression of ILDR1 AA424839 transcriptswhich are detectable by amplicon as depicted in sequence nameAA424839_seg14-16 (SEQ ID NO: 202) in different normal tissues arepresented in FIG. 20.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: AA424839_seg14-16F1R1(SEQ ID NO:202).

Forward Primer >AA424839_seg14-16F1 (SEQ ID NO: 200) GCCACCGCTACATGAAGCAReverse Primer >AA424839_seg14-16R1 (SEQ ID NO: 201) CTGGACGGCAGGGACAAATAmplicon >AA424839_seg14-16F1R1 (SEQ ID NO: 202)GCCACCGCTACATGAAGCAGGCCCAGGCCCTAGGTCCTCAGATGATGGGAAAACCCCTGTACTGGGGGGCGGACAGGAGCTCCCAGGTTTCATCTTATCCAATGCACCCGCTGCTGCAGCGAGATTTGTCCCTGCCGTCCAG

Expression of immunoglobulin-like domain containing receptor 1 (ILDR1)AA424839 transcripts which are detectable by amplicon as depicted insequence name AA424839_seg11-14F3R3 (SEQ ID NO:205) in theblood-specific panel.

Expression of ILDR1 transcripts detectable by or according toseg11-14-AA424839seg11-14F3R3 (SEQ ID NO: 205) amplicon and primersAA424839seg11-14F3 (SEQ ID NO: 203) and AA424839seg11-14R3 (SEQ ID NO:204) was measured by real time PCR on blood panel. The samples used aredetailed in Table 1 above. For each RT sample, the expression of theabove amplicon was normalized to the normalization factor calculatedfrom the expression of several house keeping genes as described inExample 1. The normalized quantity of each RT sample was then divided bythe median of the quantities of the kidney normal samples (samplenumbers 65-67, Table 1 above), to obtain a value of relative expressionof each sample relative to median of the kidney normal samples.

The results of this analysis are depicted in the histogram in FIG. 21.Expression of the above-indicated ILDR1 transcript was seen in severallymphomas and cell lines, however the expression was as high as inkidney normal samples.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: seg11-14F3 forward primer; and seg11-14R3 reverseprimer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: seg11-14F3R3.

Forward Primer >AA424839_seg11-14F3 (SEQ ID NO: 203)TCCTCCTCCTGCTGCTGATTG Reverse Primer >AA424839_seg11-14R3(SEQ ID NO: 204) TGGGCCTGCTTCATGTAGCG Amplicon >AA424839_seg11-14F3R3(SEQ ID NO: 205) TCCTCCTCCTGCTGCTGATTGGAGTGTGCTGGTGCCAGTGCTGTCCTCAGTATTGCTGCTGCTATATCCGCTGTCCCTGCTGTCCTGCCCACTGCTGCTGTCCTGAGGAAGCCCTGGCCCGCCACCGCTACATGAAGCAGGCCCA

Example 4 Description for Cluster AI216611

The present invention relates in particular to a putative B7/CD28 memberreferred to as AI216611 and diagnostics and therapeutics based thereon.According to the present invention, Cluster AI216611 (internal ID70605934) features 2 transcripts and 3 segments of interest, the namesfor which are given in Tables 49 and 50, respectively. The selectedproteins are given in table 51.

TABLE 49 Transcripts of interest Transcript Name AI216611_T0 (SEQ ID NO:41) AI216611_T1 (SEQ ID NO: 42)

TABLE 50 Segments of interest Segment Name AI216611_N2 (SEQ ID NO: 126)AI216611_N4 (SEQ ID NO: 127) AI216611_N6 (SEQ ID NO: 128)

TABLE 51 Proteins of interest Protein Name Corresponding TranscriptsAI216611_P0 (SEQ ID NO: 43) AI216611_T0 (SEQ ID NO: 41) AI216611_P1 (SEQID NO: 44) AI216611_T1 (SEQ ID NO: 42)

AI216611 is an uncharacterized gene having no full length mRNA depositedin Genbank. The protein corresponding to AI216611_P0 appears in Celera'sannotation of the human genome, based on computational analysis andtranslation of the genome (DNA sequence accession CH471065) (Venter, J.C et al., 2001 Science 291, 1304-1351). The protein corresponding toAI216611_P0 is also listed among other sequences disclosed inWO2003025148. However, this application does not characterize itsfunction or more particularly teach that it is a B7/CD28 costimulatoryprotein.

The protein corresponding to AI216611_P1 sequence is a novel protein,that is only partially similar (186 out of 199 amino acids are the same)to a polypeptide reported in WO205108415, assigned to Biogen-Idec, whichpurports that this polypeptide is a transmembrane protein that may betargeted in the treatment of hyperproliferative disorders. WO205108415does not report a function of this polypeptide. More specifically, thereis no indication that it is a B7/CD28 costimulatory protein.

According to the present invention, AI216611 is predicted to be a novelB7/CD28 family member based on the presence of an IgV domain, acharacteristic structural feature of the B7/CD28 family members. Inaddition, AI216611 is similar to the known CD28 family members in itsexons' sizes and the position of the IgV and transmembrane domainswithin these exons. Like all known B7/CD28 members, AI216611 is also atype I membrane protein. According to the present invention, twoalternatively spliced transcripts of AI216611 are provided, each onecontaining a unique region within the intracellular region. Theexpression of AI216611 and its variants was demonstrated in the presentinvention to be downregulated in colon cancer, further supporting animmune costimulatory role.

As noted above, contig AI216611 features 2 transcripts, which werelisted in Table 49 above. A description of each protein according to thepresent invention is now provided.

Protein AI216611_P0 (SEQ ID NO:43) according to the present inventionhas an amino acid sequence as encoded by transcript AI216611_T0 (SEQ IDNO:41).

The localization of the protein was determined according to results froma number of different software programs and analyses, including analysesfrom SignalP and other specialized programs. The variant protein isbelieved to be located as follows with regard to the cell: membrane.

Protein AI216611_P0 (SEQ ID NO:43) also has the following non-silentSNPs (Single Nucleotide Polymorphisms) as listed in Table 52, (givenaccording to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:43)).

TABLE 52 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 27 R −> L 27 R −> P 76 Q −> R 129 S −> R

The protein has the following domains, as determined by using InterPro.The domains are described in Table 53:

TABLE 53 InterPro domains Analysis Domain description type Positions onprotein Immunoglobulin V-set HMMPfam 33-129 domain Immunoglobulinsubtype HMMSmart 40-129

Protein AI216611_P0 (SEQ ID NO:43) is encoded by the transcriptAI216611_TO (SEQ ID NO:41), for which the coding portion starts atposition 1 and ends at position 600. The transcript also has thefollowing SNPs as listed in Table 54 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 54 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence G −> T 80, 991 G −> C 80, 991 C −> T 387, 990  C −> G 387, 1205T −> C 906, 1063, 1090 A −> G 227, 1109 T −> A 1409, 1448  G −> A 1447 T−> G 1448

The genomic structure of protein AI216611_P0 (SEQ ID NO:43) (number ofexons relevant to the extra-cellular region of the protein, the lengthof these exons, the frame of the codon in which the introns are insertedand the location of the protein features and domains in the genestructure) is characteristic to the receptors of the B7/co-stimulatoryprotein family, as given in table 55

TABLE 55 genomic structure and protein features Exon Exon Amino- numberLength Acids Protein feature on exon 1 91  1-30 Signal Peptide 2 327 31-139 Ig-like domain 3 141 140-186 Trans-membrane region 4 41 187-200

Protein AI216611_P1 (SEQ ID NO:44) according to the present inventionhas an amino acid sequence as encoded by transcript AI216611_T1 (SEQ IDNO:42).

The localization of the protein was determined according to results froma number of different software programs and analyses, including analysesfrom SignalP and other specialized programs. The variant protein isbelieved to be located as follows with regard to the cell: membrane.

Protein AI216611_P1 (SEQ ID NO:44) also has the following non-silentSNPs (Single Nucleotide Polymorphisms) as listed in Table 56, (givenaccording to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:44)).

TABLE 56 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 27 R −> L 27 R −> P 76 Q −> R 129 S −> R

The protein has the following domains, as determined by using InterPro.The domains are described in Table 57:

TABLE 57 InterPro domains Analysis Domain description type Positions onprotein Immunoglobulin V-set HMMPfam 33-129 domain Immunoglobulinsubtype HMMSmart 40-129

Protein AI216611_P1 (SEQ ID NO:44) is encoded by the transcriptAI216611_T1 (SEQ ID NO:42), for which the coding portion starts atposition 1 and ends at position 597. The transcript also has thefollowing SNPs as listed in Table 58 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 58 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence G −> T  80, 1121 G −> C  80, 1121 C −> T 387, 1120 C −> G 387,1335 T −> C 1036, 1193, 1220 A −> G 227, 1239 T −> A 1549, 1578  G −> A1577 T −> G 1578

As noted above, cluster AI216611 features 3 segments, which were listedin Table 50 above. These segments are portions of nucleic acid sequenceswhich are described herein separately because they are of particularinterest. A description of each segment according to the presentinvention is now provided.

Segment cluster AI216611_N2 (SEQ ID NO:126) according to the presentinvention is supported by 4 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: AI216611_T0 (SEQ ID NO:41) and AI216611_T1 (SEQID NO:42). Table 59 below describes the starting and ending position ofthis segment on each transcript.

TABLE 59 Segment location on transcripts Segment Segment Transcript namestarting position ending position AI216611_T0 (SEQ ID NO: 41) 92 418AI216611_T1 (SEQ ID NO: 42) 92 418

Segment cluster AI216611_N4 (SEQ ID NO:127) according to the presentinvention is supported by 4 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: AI216611_T0 (SEQ ID NO:41) and AI216611_T1 (SEQID NO:42). Table 60 below describes the starting and ending position ofthis segment on each transcript.

TABLE 60 Segment location on transcripts Segment Segment Transcript namestarting position ending position AI216611_T0 (SEQ ID NO: 41) 419 559AI216611_T1 (SEQ ID NO: 42) 419 559

Segment cluster AI216611_N6 (SEQ ID NO:128) according to the presentinvention is supported by 2 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: AI216611_T0 (SEQ ID NO:41) and AI216611_T1 (SEQID NO:42). Table 61 below describes the starting and ending position ofthis segment on each transcript.

TABLE 61 Segment location on transcripts Segment Segment Transcript namestarting position ending position AI216611_T0 (SEQ ID NO: 41) 560 885AI216611_T1 (SEQ ID NO: 42) 690 1015

Expression of AI216611 transcripts which are detectable by amplicon asdepicted in sequence name AI216611_junc4-6F2R2 (SEQ ID NO: 208) innormal and cancerous Colon tissues and in different normal tissues

Expression of AI216611 transcripts detectable by or according tojunc4-6-AI216611_junc4-6F2R2 (SEQ ID NO: 208) amplicon and primersAI216611_junc4-6F2 (SEQ ID NO: 206) and AI216611_junc4-6R2 (SEQ ID NO:207) was measured by real time PCR on colon panel and normal panel. Thesamples used are detailed in Table 5 and Table 2 above, respectively.For each RT sample, the expression of the above amplicon was normalizedto the normalization factor calculated from the expression of severalhouse keeping genes as described in Example 1.

Colon panel—The normalized quantity of each RT sample was then dividedby the median of the quantities of the normal samples (sample numbers42-70, Table 5 above). Then the reciprocal of this ratio was calculated,to obtain a value of fold down-regulation for each sample relative tomedian of the normal samples.

FIG. 22 is a histogram showing down regulation of the above-indicatedAI216611 transcripts in cancerous Colon samples relative to the normalsamples.

As is evident from FIG. 22, the expression of AI216611 transcriptsdetectable by the above amplicon in cancer samples was significantlylower than in the non-cancerous samples (sample numbers 42-70, Table 5above). Notably down regulation of at least 5 fold was found in 27 outof 55 adenocarcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. The P value for the difference in theexpression levels of AI216611 transcripts detectable by the aboveamplicon in Colon cancer samples versus the normal tissue samples wasdetermined by T test as 2.39e-005.

Threshold of 5 fold down regulation was found to differentiate betweencancer and normal samples with P value of 5.09e-007 as checked by exactFisher test.

The above values demonstrate statistical significance of the results.

Normal panel—Non-detected samples (samples no. 50, 52, 54 and 56, Table2) were assigned Ct value of 41 and were calculated accordingly. Foreach RT sample, the expression of the above amplicon was normalized tothe normalization factor calculated from the expression of several housekeeping genes as described in Example 1. The normalized quantity of eachRT sample was then divided by the median of the quantities of the colonsamples (sample numbers 3, 4 and 5, Table 2 above), to obtain a value ofrelative expression of each sample relative to median of the colonsamples, as shown in FIG. 23.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: AI216611junc4-6F2 (SEQ ID NO: 206) forward primer;and AI216611junc4-6R2 (SEQ ID NO: 207) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: AI216611 junc4-6F2R2(SEQ ID NO: 208).

Forward Primer >AI216611_junc4-6F2 (SEQ ID NO: 206):CCGCAGTATTAATCAGCCTCATG Reverse Primer >AI216611j_unc4-6R2(SEQ ID NO: 207): AATCTCCTCAGTTGTGCTTTCTTTGAmplicon >AI216611_junc4-6F2R2 (SEQ ID NO: 208)CCGCAGTATTAATCAGCCTCATGTGGGTTTGTAATAAGTGTGCATATAAATTTCAGAGGAAGAGAAGACACAAACTCAAAGAAAGCACAACTGAGG AGATT

Expression of AI216611 transcripts which are detectable by amplicon asdepicted in sequence name AI216611_seg2WT (SEQ ID NO: 211) in normal andcancerous Colon tissues and in different normal tissues

Expression of AI216611 transcripts detectable by or according toseg2WT—AI216611_seg2WT (SEQ ID NO: 211) amplicon and primersAI216611_seg2WTF1 (SEQ ID NO: 209) and AI216611_seg2WTR1 (SEQ ID NO:210) was measured by real time PCR on colon panel and normal panel. Thesamples used are detailed in Table 5 and Table 2 above, respectively.

Colon panel—Non-detected sample (sample no. 33, Table 5) was assigned Ctvalue of 41 and was calculated accordingly. For each RT sample, theexpression of the above amplicon was normalized to the normalizationfactor calculated from the expression of several house keeping genes asdescribed in Example 1. The normalized quantity of each RT sample wasthen divided by the median of the quantities of the normal samples(sample numbers 42-70, Table 5 above). Then the reciprocal of this ratiowas calculated, to obtain a value of fold down-regulation for eachsample relative to median of the normal samples.

FIG. 24 is a histogram showing down regulation of the above-indicatedAI216611 transcripts in cancerous Colon samples relative to the normalsamples.

As is evident from FIG. 24, the expression of AI216611 transcriptsdetectable by the above amplicon in cancer samples was significantlylower than in the non-cancerous samples (sample numbers 42-70, Table 5above). Notably down regulation of at least 5 fold was found in 25 outof 55 adenocarcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. Threshold of 5 fold down regulation wasfound to differentiate between cancer and normal samples with P value of2.00e-006 as checked by exact Fisher test.

The above values demonstrate statistical significance of the results.

Normal panel—For each RT sample, the expression of the above ampliconwas normalized to the normalization factor calculated from theexpression of these house keeping genes as described in Example 1,herein. The normalized quantity of each RT sample was then divided bythe median of the quantities of the colon samples (sample numbers 3, 4and 5, Table 5 above), to obtain a value of relative expression of eachsample relative to median of the colon samples, as shown in FIG. 25.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: AI216611_seg2WTF1 (SEQ ID NO: 209) forward primer;and AI216611_seg2WTR1 (SEQ ID NO: 210) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: AI216611_seg2WT (SEQID NO: 211).

Forward Primer >AI216611_seg2WTF1 (SEQ ID NO: 209) GAACGCAGAAGATCGTGGAGTReverse Primer >AI216611_seg2WTR1 (SEQ ID NO: 210) CTGAAGAGCTGGATGGAGCCAmplicon >AI216611_seg2WT (SEQ ID NO: 211)GAACGCAGAAGATCGTGGAGTGGAAACCAGGGACTCAGGCCAACATCTCTCAAAGCCACAAGGACAGAGTCTGCACCTTTGACAACGGCTCC ATCCAGCTCTTCAG

Expression of AI216611 transcripts which are detectable by amplicon asdepicted in sequence name AI216611junc4-6 (SEQ ID NO: 214) in theblood-specific panel.

Expression of AI216611 transcripts detectable by or according tojunc4-6-junc4-6F4R4 (SEQ ID NO: 214) amplicon and primers junc4-6F4 (SEQID NO: 212) and junc4-6R4 (SEQ ID NO: 213) was measured by real time PCRon blood panel. The samples used are detailed in Table 1 above. For eachRT sample, the expression of the above amplicon was normalized to thenormalization factor calculated from the expression of several housekeeping genes as described in Example 1. The normalized quantity of eachRT sample was then divided by the median of the quantities of the kidneynormal samples (sample numbers 65-67, Table 1 above), to obtain a valueof relative expression of each sample relative to median of the kidneynormal samples.

The results of this analysis are depicted in the histogram in FIG. 26.Expression of the above-indicated AI216611 transcript was much high innormal samples checked relative to the different blood specific samplesin the panel.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: junc4-6F4 (SEQ ID NO: 212) forward primer; andjunc4-6R4 reverse primer (SEQ ID NO: 213).

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: junc4-6F4R4 (SEQ IDNO: 214).

Forward Primer AI216611junc4-6F4 (SEQ ID NO:  212):CTGCACTTTGTCGCTGTCATC Reverse Primer AI216611junc4-6R4 (SEQ ID NO: 213):CAATCTCCTCAGTTGTGCTTTCTTTGAmplicon AI216611junc4-6F4R4 (SEQ ID NO: 214):CTGCACTTTGTCGCTGTCATCCTTGCTTTTCTCGCTGCTGTGGCCGCAGTATTAATCAGCCTCATGTGGGTTTGTAATAAGTGTGCATATAAATTTCAGAGGAAGAGAAGACACAAACTCAAAGAAAGCACAACTGAGGAGATTG

Expression of AI216611 transcripts which are detectable by amplicon asdepicted in sequence name AI216611junc2-4seg5 (SEQ ID NO: 217) in theblood-specific panel.

Expression of AI216611 transcripts detectable by or according tojunc2-4seg5-junc2-4seg5F3R4 amplicon (SEQ ID NO: 217) and primersjunc2-4seg5F3 (SEQ ID NO: 215) and junc2-4seg5R4 (SEQ ID NO: 216) wasmeasured by real time PCR on blood panel. The samples used are detailedin Table 1 above. For each RT sample, the expression of the aboveamplicon was normalized to the normalization factor calculated from theexpression of several house keeping genes as described in Example 1. Thenormalized quantity of each RT sample was then divided by the median ofthe quantities of the kidney normal samples (sample numbers 65-67, Table1 above), to obtain a value of relative expression of each samplerelative to median of the kidney normal samples.

The results of this analysis are depicted in the histogram in FIG. 27.Expression of the above-indicated AI216611 transcript was much high innormal samples checked relative to the different blood specific samplesin the panel.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: junc2-4seg5F3 (SEQ ID NO: 215) forward primer; andjunc2-4seg5R4 (SEQ ID NO: 216) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: junc2-4seg5F3R4 (SEQID NO: 217).

Forward Primer AI216611 junc2-4seg5F3 (SEQ ID NO: 215):TGCTGCACGTCTCTGAGATCC Reverse Primer AI216611 junc2-4seg5R4(SEQ ID NO: 216): CACCTCTGGCCTCAAAACCACTCAmplicon >AI216611 junc2-4seg5F3R4 (SEQ ID NO: 217)TGCTGCACGTCTCTGAGATCCTCTATGAAGACCTGCACTTTGTCGCTGTCATCCTTGCTTTTCTCGCTGCTGTGGCCGCAGTATTAATCAGCCTCATGTGGGTTTGTAATAAGTGTGCATATAAATTTCAGAGGAAGAGAAGACACAAACTCAAAGGTAACCCCCTGGGCCTTGTGATAATCCATGAGTGGTTTTG AGGCCAGAGGTG

Expression of AI216611 transcripts which are detectable by amplicon asdepicted in sequence name AI216611_junc2-4seg5F2R2 (SEQ ID NO: 220) innormal and cancerous Colon tissues

Expression of AI216611 transcripts detectable by or according tojunc2-4seg5F2R2—AI216611_junc2-4seg5F2R2 (SEQ ID NO: 220) amplicon andprimers AI216611_junc2-4seg5F2 (SEQ ID NO: 218) andAI216611_junc2-4seg5R2 (SEQ ID NO: 219) was measured by real time PCR oncolon panel. The samples used are detailed in Table 5 above.Non-detected samples (samples no. 28, 33, 83, 85, 90 and 63, Table 5)were assigned Ct value of 41 and were calculated accordingly. For eachRT sample, the expression of the above amplicon was normalized to thenormalization factor calculated from the expression of several housekeeping genes as described in Example 1. The normalized quantity of eachRT sample was then divided by the median of the quantities of the normalsamples (sample numbers 42-62 and 64-70, Table 5 above). Then thereciprocal of this ratio was calculated, to obtain a value of folddown-regulation for each sample relative to median of the normalsamples.

FIG. 28 is a histogram showing down regulation of the above-indicatedAI216611 transcripts in cancerous Colon samples relative to the normalsamples.

As is evident from FIG. 28, the expression of AI216611 transcriptsdetectable by the above amplicon in cancer samples was significantlylower than in the non-cancerous samples (sample numbers 42-62 and 64-70,Table 5 above). Notably down regulation of at least 5 fold was found in31 out of 55 adenocarcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. The P value for the difference in theexpression levels of AI216611 transcripts detectable by the aboveamplicon in Colon cancer samples versus the normal tissue samples wasdetermined by T test as 5.29e-003.

Threshold of 5 fold down regulation was found to differentiate betweencancer and normal samples with P value of 4.18e-008 as checked by exactFisher test.

The above values demonstrate statistical significance of the results.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: AI216611_junc2-4seg5F2 (SEQ ID NO: 218) forwardprimer; and AI216611_junc2-4seg5R2 (SEQ ID NO: 219) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon:AI216611_junc2-4seg5F2R2 (SEQ ID NO: 220).

Forward Primer >AI216611_junc2-4seg5F2 (SEQ ID NO: 218)GCTGCACGTCTCTGAGATCCT Reverse Primer >AI216611_junc2-4seg5R2(SEQ ID NO: 219) CACCTCTGGCCTCAAAACCA Amplicon >AI216611_junc2-4seg5F2R2(SEQ ID NO: 220) GCTGCACGTCTCTGAGATCCTCTATGAAGACCTGCACTTTGTCGCTGTCATCCTTGCTTTTCTCGCTGCTGTGGCCGCAGTATTAATCAGCCTCATGTGGGTTTGTAATAAGTGTGCATATAAATTTCAGAGGAAGAGAAGACACAAACTCAAAGGTAACCCCCTGGGCCTTGTGATAATCCATGAGTGGTTTTGA GGCCAGAGGTG

Example 5 Description for Cluster H68654_(—)1

The present invention relates to LOC253012 polypeptides, novel splicevariants and diagnostics and therapeutics based thereon.

According to the present invention, Cluster H68654_(—)1 (internal ID76432882) features 10 transcripts and 3 segments of interest, the namesfor which are given in Tables 62 and 63, respectively. The selectedprotein variants are given in table 64.

TABLE 62 Transcripts of interest Transcript Name H68654_1_T0 (SEQ ID NO:25) H68654_1_T4 (SEQ ID NO: 26) H68654_1_T5 (SEQ ID NO: 27) H68654_1_T8(SEQ ID NO: 28) H68654_1_T15 (SEQ ID NO: 29) H68654_1_T16 (SEQ ID NO:30) H68654_1_T17 (SEQ ID NO: 31) H68654_1_T18 (SEQ ID NO: 32)H68654_1_T19 (SEQ ID NO: 33) H68654_1_T20 (SEQ ID NO: 34)

TABLE 63 Segments of interest Segment Name H68654_1_N3 (SEQ ID NO: 123)H68654_1_N7 (SEQ ID NO: 124) H68654_1_N12 (SEQ ID NO: 125)

TABLE 64 Proteins of interest Protein Name Corresponding TranscriptsH68654_1_P2 (SEQ ID NO: 35) H68654_1_T0 (SEQ ID NO: 25); H68654_1_T5(SEQ ID NO: 27) H68654_1_P5 (SEQ ID NO: 36) H68654_1_T4 (SEQ ID NO: 26)H68654_1_P7 (SEQ ID NO: 37) H68654_1_T8 (SEQ ID NO: 28) H68654_1_P12(SEQ ID NO: 38) H68654_1_T15 (SEQ ID NO: 29); H68654_1_T16 (SEQ ID NO:30); H68654_1_T18 (SEQ ID NO: 32) H68654_1_P13 (SEQ ID NO: 39)H68654_1_T17 (SEQ ID NO: 31); H68654_1_T19 (SEQ ID NO: 33) H68654_1_P14(SEQ ID NO: 40) H68654_1_T20 (SEQ ID NO: 34)

These sequences are variants of the known protein hypothetical proteinLOC253012 isoform 1 (RefSeq accession identifier NP_(—)001034461 (SEQ IDNO: 35), NP_(—)937794 (SEQ ID NO: 36)), referred to herein as thepreviously known protein.

The known LOC253012 is a hypothetical protein that was computationallydiscovered during the secreted protein discovery initiative project(SPDI), a large-scale effort to identify novel human secreted andtransmembrane proteins (Clark et al 2003, Genome Res. 13: 2265-2270).Its closest paralog that is experimentally validated is the hepatocytecell adhesion molecule (Refseq accession NP_(—)689935) (evalue e-21).

LOC253012 antigen has been reported in European patent applicationNumber EP162070, assigned to Genentech Inc. purports that LOC253012(PRO346), corresponding to H68654_(—)1_P2 (SEQ ID NO:35), isdifferentially expressed in a lung tumor samples. This patentspecification alleges that the corresponding polypeptide can be used fordeveloping antibodies against the disclosed PRO antigens, includingPRO346. The patent application further suggests that antibodies to agenus of polypeptides reported therein that includes PRO346 may be usedfor treating and diagnosing cancer and specifically for diagnosing andtreating lung cancer. The Genentech patent application does not teach,however, that LOC253012 (PRO346) is differentially expressed in smallcell lung carcinoma. Also, there is no specific teaching in GenentechInc. patent application that the anti-PRO346 antibodies can be used fortreating small cell lung carcinomas and/or for modulating co-stimulationof the APC/T cell activity. Also, there is no teaching in the GenentechInc. patent application that the PRO346 is an immune costimulatoryprotein or more specifically a B7 family member. There is no teaching inthe Genentech Inc. patent application of the use of antibodies againstthe PRO346 antigen for modulating immune co-stimulation or particularlythe B7 co-stimulatory pathway.

According to the present invention, LOC253012 was predicted to be anovel immune costimulatory protein and in particular a B7 co-stimulatoryprotein. The prediction was based on the presence of both an IgV domainand IgC2, a characteristic structural feature of the B7-family members.Like other B7 members, LOC253012 is also a type I membrane protein.LOC253012 and its variants were demonstrated in the present invention tobe overexpressed in lung cancer.

MED discovery engine described in Example 1 herein, was used to assessthe expression of LOC253012 transcripts. Expression data for Affymetrixprobe set 242601_at representing the LOC253012 gene data is shown inFIG. 29. As evident from the scatter plot, presented in FIG. 29, theexpression of LOC253012 transcripts detectable with the above probe setswas higher in lung cancer compared to normal lung samples.

As noted above, cluster H68654 features 10 transcripts, which werelisted in Table 62 above. These transcripts encode for proteins whichare variants of protein hypothetical protein LOC253012 isoform 1 (SEQ IDNO:35). A description of each variant protein according to the presentinvention is now provided.

Variant protein H68654_(—)1_P2 (SEQ ID NO:35) according to the presentinvention has an amino acid sequence as encoded by transcriptsH68654_(—)1_T0 (SEQ ID NO:25) and H68654_(—)1_T5 (SEQ ID NO:27).

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein H68654_(—)1_P2 (SEQ ID NO:35) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 65,(given according to their positions on the amino acid sequence, with thealternative amino acids listed).

TABLE 65 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 33 K −> R 86 K −> N 204 L −> P

The variant protein has the following domains, as determined by usingInterPro. The domains are described in Table 66:

TABLE 66 InterPro domains Analysis Domain description type Positions onprotein Immunoglobulin subtype HMMSmart 38-141, 155-235, 255-333Immunoglobulin-like ProfileScan 149-233, 235-331 Immunoglobulin-likeHMMPfam 163-221 Immunoglobulin V-set HMMPfam  31-143 Immunoglobulin C2type HMMSmart 164-226

The coding portion of transcript H68654_(—)1_T0 (SEQ ID NO:25) starts atposition 79 and ends at position 1464. The transcript also has thefollowing SNPs as listed in Table 67 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 67 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 176, 1545 G −> A 336, 1754 G −> C  336 T −> C 564, 689 A −> T 1545 G −> T 1684 T −> A 1755 T −> G 1755

The coding portion of transcript H68654_(—)1_T5 (SEQ ID NO:27) starts atposition 79 and ends at position 1464. The transcript also has thefollowing SNPs as listed in Table 68 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 68 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 176, 1545 G −> A 336, 1754 G −> C  336 T −> C 564, 689 A −> T 1545 G −> T 1684 T −> A 1755 T −> G 1755

The genomic structure of protein H68654_(—)1_P2 (SEQ ID NO: 35) (numberof exons relevant to the extra-cellular region of the protein, thelength of these exons, the frame of the codon in which the introns areinserted and the location of the protein features and domains in thegene structure) is characteristic to the ligands of theB7/co-stimulatory protein family, as given in table 69.

TABLE 69 genomic structure and protein features Exon Exon Amino- numberLength Acids Protein feature on exon 1 79  1-26 Signal Peptide 2 351 27-143 IgV domain 3 285 144-238 IgC2 domain 4 297 239-337 Ig-likedomain 5 126 338-379 Trans-membrane region 6 25 380-387 7 38 388-400 874 401-425 9 110 426-462 10 1 462-462

Variant protein H68654_(—)1_P5 (SEQ ID NO:36) according to the presentinvention has an amino acid sequence encoded by transcriptsH68654_(—)1_T4 (SEQ ID NO:26).

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein H68654_(—)1_P5 (SEQ ID NO:36) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 70,(given according to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:36)).

TABLE 70 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 21 K −> R 74 K −> N 192 L −> P

The variant protein has the following domains, as determined by usingInterPro. The domains are described in Table 71:

TABLE 71 InterPro domains Analysis Domain description type Positions onprotein Immunoglobulin C2 type HMMSmart 152-214 Immunoglobulin subtypeHMMSmart 26-129, 143-223, 243-321 Immunoglobulin-like HMMPfam 151-209Immunoglobulin V-set HMMPfam  19-131 Immunoglobulin-like Profile137-221, 223-319

Variant protein H68654_(—)1_P5 (SEQ ID NO:36) is encoded by thetranscript H68654_(—)1_T4 (SEQ ID NO:26), for which the coding portionstarts at position 102 and ends at position 1451. The transcript alsohas the following SNPs as listed in Table 72 (given according to theirposition on the nucleotide sequence, with the alternative nucleic acidlisted).

TABLE 72 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 163, 1532 G −> A 323, 1741 G −> C  323 T −> C 551, 676 A −> T 1532 G −> T 1671 T −> A 1742 T −> G 1742

Variant protein H68654_(—)1_P7 (SEQ ID NO:37) according to the presentinvention has an amino acid sequence encoded by transcriptH68654_(—)1_T8 (SEQ ID NO:28). Alignment of H68654_(—)1_P7 (SEQ IDNO:37) to one or more previously published protein sequences are shownin FIGS. 30A and 30B. A brief description of the relationship of thevariant protein according to the present invention to each such alignedprotein is as follows:

1. Comparison report between H68654_(—)1_P7 (SEQ ID NO:37) and knownproteins NP_(—)937794 and Q6UXI0_HUMAN (SEQ ID NO: 36) (FIG. 30A):

A. An isolated chimeric polypeptide encoding for H68654_(—)1_P7 (SEQ IDNO:37), comprising a first amino acid sequence being at least 90%homologous to MWLKVFTTFLSFATGACSGLKVTVPSHTVHGVRGQALYLPVHYGFHTPASDIQIIWLFERPHTMPKYLLGSVNKSVVPDLEYQHKFTMMPPNASLLINPLQFPDEGNYIVKVNIQGNGTLSASQKIQVTVDDPVTKPVVQIHPPSGAVEYVGNMTLTCHVEGGTRLAYQWLKNGRPVHTSSTYSFSPQNNTLHIAPVTKEDIGNYSCLVRNPVSEMESDIIMPIIYYGPYGLQVNSDKGLKVGEVFTVDLGEAILFDCSADSHPPNTYSWIRRTDNTTYIIKHGPRLEVASEKVAQKTMDYVCCAYNNITGRQDETHFTVIITSVGLEKLAQKGKSLSPLASITGISLFLIISMCLLFLWKKYQPYK corresponding to amino acids1-367 of known proteins NP_(—)937794 and Q6UXI0_HUMAN (SEQ ID NO: 36),which also corresponds to amino acids 1-367 of H68654_(—)1_P7 (SEQ IDNO:37), and a second amino acid sequence being at least 70%, optionallyat least 80%, preferably at least 85%, more preferably at least 90% andmost preferably at least 95, 96, 97, 98 or 99% homologous to apolypeptide having the sequenceGQKQNTGKLKHFQAMKMLWMTSEYMNLLLFQMFLVFPGSQAGLFQPLIVYRGKICTVQCMKLFSTSLPSSKTIQSELSWAKQYIRVKF (SEQ ID NO: 290) corresponding toamino acids 368-455 of H68654_(—)1_P7 (SEQ ID NO:37), wherein said firstamino acid sequence and second amino acid sequence are contiguous and ina sequential order.

B. An isolated polypeptide encoding for an edge portion ofH68654_(—)1_P7 (SEQ ID NO:37), comprising an amino acid sequence beingat least 70%, optionally at least about 80%, preferably at least about85%, more preferably at least about 90% and most preferably at leastabout 95, 96, 97, 98 or 99% homologous to the sequenceGQKQNTGKLKHFQAMKMLWMTSEYMNLLLFQMFLVFPGSQAGLFQPLIVYRGKICTVQCMKLFSTSLPSSKTIQSELSWAKQYIRVKF (SEQ ID NO: 290) of H68654_(—)1_P7(SEQ ID NO:37).

2. Comparison report between H68654_(—)1_P7 (SEQ ID NO:37) and knownproteins NP_(—)001034461 (SEQ ID NO: 35) FIG. 30B):

A. An isolated chimeric polypeptide encoding for H68654_(—)1_P7 (SEQ IDNO:37), comprising a first amino acid sequence being at least 70%,optionally at least 80%, preferably at least 85%, more preferably atleast 90% and most preferably at least 95, 96, 97, 98 or 99%, homologousto a polypeptide having the sequence MWLKVFTTFLSFAT (SEQ ID NO: 289)corresponding to amino acids 1-14 of H68654_(—)1_P7 (SEQ ID NO:37), asecond amino acid sequence being at least 90% homologous toGACSGLKVTVPSHTVHGVRGQALYLPVHYGFHTPASDIQIIWLFERPHTMPKYLLGSVNKSVVPDLEYQHKFTMMPPNASLLINPLQFPDEGNYIVKVNIQGNGTLSASQKIQVTVDDPVTKPVVQIHPPSGAVEYVGNMTLTCHVEGGTRLAYQWLKNGRPVHTSSTYSFSPQNNTLHIAPVTKEDIGNYSCLVRNPVSEMESDIIMPIIYYGPYGLQVNSDKGLKVGEVFTVDLGEAILFDCSADSHPPNTYSWIRRTDNTTYIIKHGPRLEVASEKVAQKTMDYVCCAYNNITGRQDETHFTVIITSVGLEKLAQKGKSLSPLASITGISLFLIISMCLLFLWKKYQPYK corresponding to amino acids 27-379 of knownproteins NP_(—)001034461 (SEQ ID NO: 35), which also corresponds toamino acids 15-367 of H68654_(—)1_P7 (SEQ ID NO:37), and a third aminoacid sequence being at least 70%, optionally at least 80%, preferably atleast 85%, more preferably at least 90% and most preferably at least 95,96, 97, 98 or 99% homologous to a polypeptide having the sequenceGQKQNTGKLKHFQAMKMLWMTSEYMNLLLFQMFLVFPGSQAGLFQPLIVYRGKICTVQCMKLFSTSLPSSKTIQSELSWAKQYIRVKF (SEQ ID NO: 290) corresponding toamino acids 368-455 of H68654_(—)1_P7 (SEQ ID NO:37), wherein said firstamino acid sequence, second amino acid sequence and third amino acidsequence are contiguous and in a sequential order.

B. An isolated polypeptide encoding for a head of H68654_(—)1_P7 (SEQ IDNO:37), comprising a polypeptide being at least 70%, optionally at leastabout 80%, preferably at least about 85%, more preferably at least about90% and most preferably at least about 95, 96, 97, 98 or 99% homologousto the sequence MWLKVFTTFLSFAT (SEQ ID NO: 289) of H68654_(—)1_P7 (SEQID NO:37).

C. An isolated polypeptide encoding for an edge portion ofH68654_(—)1_P7 (SEQ ID NO:37), comprising an amino acid sequence beingat least 70%, optionally at least about 80%, preferably at least about85%, more preferably at least about 90% and most preferably at leastabout 95, 96, 97, 98 or 99% homologous to the sequenceGQKQNTGKLKHFQAMKMLWMTSEYMNLLLFQMFLVFPGSQAGLFQPLIVYRGKICTVQCMKLFSTSLPSSKTIQSELSWAKQYIRVKF (SEQ ID NO: 290) of H68654_(—)1_P7(SEQ ID NO:37).

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein H68654_(—)1_P7 (SEQ ID NO:37) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 73,(given according to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:37)).

TABLE 73 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 21 K −> R 74 K −> N 192 L −> P

The variant protein has the following domains, as determined by usingInterPro. The domains are described in Table 74:

TABLE 74 InterPro domains Analysis Domain description type Positions onprotein Immunoglobulin subtype HMMSmart 26-129, 143-223, 243-321Immunoglobulin C2 type HMMSmart 152-214 Immunoglobulin-like ProfileScan137-221, 223-319 Immunoglobulin V-set HMMPfam  19-131Immunoglobulin-like HMMPfam 151-209

Variant protein H68654_(—)1_P7 (SEQ ID NO:37) is encoded by thetranscript H68654_(—)1_T8 (SEQ ID NO:28), for which the coding portionstarts at position 102 and ends at position 1466. The transcript alsohas the following SNPs as listed in Table 75 (given according to theirposition on the nucleotide sequence, with the alternative nucleic acidlisted).

TABLE 75 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 163, 1507 G −> A 323, 1716 G −> C  323 T −> C 551, 676 A −> T 1507 G −> T 1646 T −> A 1717 T −> G 1717

Variant protein H68654_(—)1_P12 (SEQ ID NO:38) according to the presentinvention has an amino acid sequence as encoded by transcriptsH68654_(—)1_T15 (SEQ ID NO:29), H68654_(—)1_T16 (SEQ ID NO:30) andH68654_(—)1_T18 (SEQ ID NO:32). Alignment of H68654_(—)1_P12 (SEQ IDNO:38) to previously published protein sequences are shown in FIGS. 30Cand 30D. A brief description of the relationship of the variant proteinaccording to the present invention to each such aligned protein is asfollows:

1. Comparison report between H68654_(—)1_P12 (SEQ ID NO:38) and knownproteins NP_(—)937794 and Q6UXI0_HUMAN (SEQ ID NO: 36) (FIG. 30C):

A. An isolated chimeric polypeptide encoding for H68654_(—)1_P12 (SEQ IDNO:38), comprising a first amino acid sequence being at least 90%homologous to MWLKVFTTFLSFATGACSGLKVTVPSHTVHGVRGQALYLPVHYGFHTPASDIQIIWLFERPHTMPKYLLGSVNKSVVPDLEYQHKFTMMPPNASLLINPLQFPDEGNYIVKVNIQGNGTLSASQKIQVTVDDPVTKPVVQIHPPSGAVEYVGNMTLTCHVEGGTRLAYQWLKNGRPVHTSSTYSFSPQNNTLHIAPVTKEDIGNYSCLVRNPVSEMESDIIMPIIYYGPYGLQVNSDKGLKVGEVFTVDLGEAILFDCSADSHPPNTYSWIRRTDNTTYIIKHGPRLEVASEKVAQKTMDYVCCAYNNITGRQDETHFTVIITSVGLEKLAQKGKSLSPLASITGISLFLIISMCLLFLWKKYQPYKVIKQKLEGRPETEYRKAQTFSGHEDALDDFGIYEFVAFPDVSGVSR corresponding to amino acids 1-413 ofknown proteins NP_(—)937794 and Q6UXI0_HUMAN (SEQ ID NO: 36), which alsocorresponds to amino acids 1-413 of H68654_(—)1_P12 (SEQ ID NO:38), anda second amino acid sequence being at least 70%, optionally at least80%, preferably at least 85%, more preferably at least 90% and mostpreferably at least 95, 96, 97, 98 or 99% homologous to a polypeptidehaving the sequence VGFPSG (SEQ ID NO: 291) corresponding to amino acids414-419 of H68654_(—)1_P12 (SEQ ID NO:38), wherein said first amino acidsequence and second amino acid sequence are contiguous and in asequential order.

B. An isolated polypeptide encoding for an edge portion ofH68654_(—)1_P12 (SEQ ID NO:38), comprising an amino acid sequence beingat least 70%, optionally at least about 80%, preferably at least about85%, more preferably at least about 90% and most preferably at leastabout 95, 96, 97, 98 or 99% homologous to the sequence VGFPSG (SEQ IDNO: 291) of H68654_(—)1_P12 (SEQ ID NO:38).

2. Comparison report between H68654_(—)1_P12 (SEQ ID NO:38) and knownproteins NP_(—)001034461 (SEQ ID NO: 35) (FIG. 30D):

A. An isolated chimeric polypeptide encoding for H68654_(—)1_P12 (SEQ IDNO:38), comprising a first amino acid sequence being at least 70%,optionally at least 80%, preferably at least 85%, more preferably atleast 90% and most preferably at least 95, 96, 97, 98 or 99%, homologousto a polypeptide having the sequence MWLKVFTTFLSFAT (SEQ ID NO: 289)corresponding to amino acids 1-14 of H68654_(—)1_P12 (SEQ ID NO:38), asecond amino acid sequence being at least 90% homologous toGACSGLKVTVPSHTVHGVRGQALYLPVHYGFHTPASDIQIIWLFERPHTMPKYLLGSVNKSVVPDLEYQHKFTMMPPNASLLINPLQFPDEGNYIVKVNIQGNGTLSASQKIQVTVDDPVTKPVVQIHPPSGAVEYVGNMTLTCHVEGGTRLAYQWLKNGRPVHTSSTYSFSPQNNTLHIAPVTKEDIGNYSCLVRNPVSEMESDIIMPIIYYGPYGLQVNSDKGLKVGEVFTVDLGEAILFDCSADSHPPNTYSWIRRTDNTTYIIKHGPRLEVASEKVAQKTMDYVCCAYNNITGRQDETHFTVIITSVGLEKLAQKGKSLSPLASITGISLFLIISMCLLFLWKKYQPYKVIKQKLEGRPETEYRKAQTFSGHEDALDDFGI YEFVAFPDVSGVSRcorresponding to amino acids 27-425 of known proteins NP_(—)001034461(SEQ ID NO: 35), which also corresponds to amino acids 15-413 ofH68654_(—)1_P12 (SEQ ID NO:38), and a third amino acid sequence being atleast 70%, optionally at least 80%, preferably at least 85%, morepreferably at least 90% and most preferably at least 95, 96, 97, 98 or99% homologous to a polypeptide having the sequence VGFPSG (SEQ ID NO:291) corresponding to amino acids 414-419 of H68654_(—)1_P12 (SEQ IDNO:38), wherein said first amino acid sequence, second amino acidsequence and third amino acid sequence are contiguous and in asequential order.

B. An isolated polypeptide encoding for a head of H68654_(—)1_P12 (SEQID NO:38), comprising a polypeptide being at least 70%, optionally atleast about 80%, preferably at least about 85%, more preferably at leastabout 90% and most preferably at least about 95, 96, 97, 98 or 99%homologous to the sequence MWLKVFTTFLSFAT (SEQ ID NO: 289) ofH68654_(—)1_P12 (SEQ ID NO:38).

C. An isolated polypeptide encoding for an edge portion ofH68654_(—)1_P12 (SEQ ID NO:38), comprising an amino acid sequence beingat least 70%, optionally at least about 80%, preferably at least about85%, more preferably at least about 90% and most preferably at leastabout 95, 96, 97, 98 or 99% homologous to the sequence VGFPSG (SEQ IDNO: 291) of H68654_(—)1_P12 (SEQ ID NO:38).

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein H68654_(—)1_P12 (SEQ ID NO:38) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 76,(given according to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:38)).

TABLE 76 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 21 K −> R 74 K −> N 192 L −> P

The variant protein has the following domains, as determined by usingInterPro. The domains are described in Table 77.

TABLE 77 InterPro domains Analysis Domain description type Positions onprotein Immunoglobulin subtype HMMSmart 26-129, 143-223, 243-321Immunoglobulin-like HMMPfam 151-209 Immunoglobulin C2 type HMMSmart152-214 Immunoglobulin V-set HMMPfam  19-131 Immunoglobulin-likeProfileScan 137-221, 223-319

The coding portion of transcript H68654_(—)1_T15 (SEQ ID NO:29) startsat position 102 and ends at position 1358. The transcript also has thefollowing SNPs as listed in Table 78 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 78 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 163, 1586 G −> A 323, 1795 G −> C  323 T −> C 551, 676 A −> T 1586 G −> T 1725 T −> A 1796 T −> G 1796

The coding portion of transcript H68654_(—)1_ starts at position 102 andends at position 1358. The transcript also has the following SNPs aslisted in Table 79 (given according to their position on the nucleotidesequence, with the alternative nucleic acid listed).

TABLE 79 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 163, 1588 G −> A 323, 1797 G −> C  323 T −> C 551, 676 A −> T 1588 G −> T 1727 T −> A 1798 T −> G 1798

The coding portion of transcript H68654_(—)1_T18 (SEQ ID NO:32) startsat position 102 and ends at position 1358. The transcript also has thefollowing SNPs as listed in Table 80 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 80 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G  163 G −> A 323, 2117 G −> C  323 T −> C 551, 676  T −>A 1603 T −> G 1603

Variant protein H68654_(—)1_P13 (SEQ ID NO:39) according to the presentinvention has an amino acid sequence as encoded by transcriptsH68654_(—)1_T17 (SEQ ID NO:31) and H68654_(—)1_T19 (SEQ ID NO:33).Alignments to one or more previously published protein sequences areshown in FIGS. 30E and 30F. A brief description of the relationship ofthe variant protein according to the present invention to each suchaligned protein is as follows:

1. Comparison report between H68654_(—)1_P13 (SEQ ID NO:39) and knownproteins NP_(—)937794 and Q6UXI0_HUMAN (SEQ ID NO: 36) (FIG. 30E):

A. An isolated chimeric polypeptide encoding for H68654_(—)1_P13 (SEQ IDNO:39), comprising a amino acid sequence being at least 90% homologousto MWLKVFTTFLSFATGACSGLKVTVPSHTVHGVRGQALYLPVHYGFHTPASDIQIIWLFERPHTMPKYLLGSVNKSVVPDLEYQHKFTMMPPNASLLINPLQFPDEGNYIVKVNIQGNGTLSASQKIQVTVDDPVTKPVVQIHPPSGAVEYVGNMTLTCHVEGGTRLAYQWLKNGRPVHTSSTYSFSPQNNTLHIAPVTKEDIGNYSCLVRNPVSEMESDIIMPIIYYGPYGLQVNSDKGLKVGEVFTVDLGEAILFDCSADSHPPNTYSWIRRTDNTTYIIKHGPRLEVASEKVAQKTMDYVCCAYNNITGRQDETHFTVIITSVGLEKLAQKGKSLSPLASITGISLFLIISMCLLFLWKKYQPYKVIKQKLEGR corresponding to aminoacids 1-376 of known proteins NP_(—)937794 and Q6UXI0_HUMAN (SEQ ID NO:36), which also corresponds to amino acids 1-376 of H68654_(—)1_P13 (SEQID NO:39), wherein said and first amino acid sequence are contiguous andin a sequential order.

2. Comparison report between H68654_(—)1_P13 (SEQ ID NO:39) and knownproteins NP_(—)001034461 (SEQ ID NO: 35) (FIG. 30F):

A. An isolated chimeric polypeptide encoding for H68654_(—)1_P13 (SEQ IDNO:39), comprising a first amino acid sequence being at least 70%,optionally at least 80%, preferably at least 85%, more preferably atleast 90% and most preferably at least 95, 96, 97, 98 or 99%, homologousto a polypeptide having the sequence MWLKVFTTFLSFAT (SEQ ID NO: 289)corresponding to amino acids 1-14 of H68654_(—)1_P13 (SEQ ID NO:39), anda second amino acid sequence being at least 90% homologous toGACSGLKVTVPSHTVHGVRGQALYLPVHYGFHTPASDIQIIWLFERPHTMPKYLLGSVNKSVVPDLEYQHKFTMMPPNASLLINPLQFPDEGNYIVKVNIQGNGTLSASQKIQVTVDDPVTKPVVQIHPPSGAVEYVGNMTLTCHVEGGTRLAYQWLKNGRPVHTSSTYSFSPQNNTLHIAPVTKEDIGNYSCLVRNPVSEMESDIIMPIIYYGPYGLQVNSDKGLKVGEVFTVDLGEAILFDCSADSHPPNTYSWIRRTDNTTYIIKHGPRLEVASEKVAQKTMDYVCCAYNNITGRQDETHFTVIITSVGLEKLAQKGKSLSPLASITGISLFLIISMCLLFLWKKYQPYKVIKQKLEGR corresponding to amino acids 27-388of known proteins NP_(—)001034461 (SEQ ID NO: 35), which alsocorresponds to amino acids 15-376 of H68654_(—)1_P13 (SEQ ID NO:39),wherein said first amino acid sequence and second amino acid sequenceare contiguous and in a sequential order.

B. An isolated polypeptide encoding for a head of H68654_(—)1_P13 (SEQID NO:39), comprising a polypeptide being at least 70%, optionally atleast about 80%, preferably at least about 85%, more preferably at leastabout 90% and most preferably at least about 95, 96, 97, 98 or 99%homologous to the sequence MWLKVFTTFLSFAT (SEQ ID NO: 289) ofH68654_(—)1_P13 (SEQ ID NO:39).

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein H68654_(—)1_P13 (SEQ ID NO:39) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 81,(given according to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:39)).

TABLE 81 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 21 K −> R 74 K −> N 192 L −> P

The variant protein has the following domains, as determined by usingInterPro. The domains are described in Table 82:

TABLE 82 InterPro domains Analysis Domain description type Positions onprotein Immunoglobulin subtype HMMSmart 26-129, 143-223, 243-321Immunoglobulin-like ProfileScan 137-221, 223-319 Immunoglobulin C2 typeHMMSmart 152-214 Immunoglobulin V-set HMMPfam  19-131Immunoglobulin-like HMMPfam 151-209

The coding portion of transcript H68654_(—)1_T17 (SEQ ID NO:31) startsat position 102 and ends at position 1229. The transcript also has thefollowing SNPs as listed in Table 83 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 83 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 163, 1626 G −> A 323, 1835 G −> C  323 T −> C 551, 676 A −> T 1626 G −> T 1765 T −> A 1836 T −> G 1836

The coding portion of transcript H68654_(—)1_T19 (SEQ ID NO:33) startsat position 102 and ends at position 1229. The transcript also has thefollowing SNPs as listed in Table 84 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 84 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G  163 G −> A 323, 2211 G −> C  323 T −> C 551, 676  T −>A 1697 T −> G 1697

Variant protein H68654_(—)1_P14 (SEQ ID NO:40) according to the presentinvention has an amino acid sequence as encoded by transcriptH68654_(—)1_T20 (SEQ ID NO:34). Alignments to previously publishedprotein sequences are shown in FIGS. 30G and 30H. A brief description ofthe relationship of the variant protein according to the presentinvention to each such aligned protein is as follows:

1. Comparison report between H68654_(—)1_P14 (SEQ ID NO:40) and knownproteins NP_(—)937794 and Q6UXI0_HUMAN (SEQ ID NO: 36) (FIG. 30G):

A. An isolated chimeric polypeptide encoding for H68654_(—)1_P14 (SEQ IDNO:40), comprising a first amino acid sequence being at least 90%homologous to MWLKVFTTFLSFATGACSGLKVTVPSHTVHGVRGQALYLPVHYGFHTPASDIQIIWLFERPHTMPKYLLGSVNKSVVPDLEYQHKFTMMPPNASLLINPLQFPDEGNYIVKVNIQGNGTLSASQKIQVTVDDPVTKPVVQIHPPSGAVEYVGNMTLTCHVEGGTRLAYQWLKNGRPVHTSSTYSFSPQNNTLHIAPVTKEDIGNYSCLVRNPVSEMESDIIMPIIYYGPYGLQVNSDKGLKVGEVFTVDLGEAILFDCSADSHPPNTYSWIRRTDNTTYIIKHGPRLEVASEKVAQKTMDYVCCAYNNITGRQDETHFTVIITSVGLEKLAQKGKSLSPLASITGISLFLIISMCLLFLWKKYQPYKVIKQKLEGRPETEYRKA QTFSGcorresponding to amino acids 1-389 of known proteins NP_(—)937794 andQ6UXI0_HUMAN (SEQ ID NO: 36), which also corresponds to amino acids1-389 of H68654_(—)1_P14 (SEQ ID NO:40), and a second amino acidsequence being at least 70%, optionally at least 80%, preferably atleast 85%, more preferably at least 90% and most preferably at least 95,96, 97, 98 or 99% homologous to a polypeptide having the sequenceFMLAAPSQREEEKKIWQGPGLLLCPHCNPHYHQY (SEQ ID NO: 292) corresponding toamino acids 390-423 of H68654_(—)1_P14 (SEQ ID NO:40), wherein saidfirst amino acid sequence and second amino acid sequence are contiguousand in a sequential order.

B. An isolated polypeptide encoding for an edge portion ofH68654_(—)1_P14 (SEQ ID NO:40), comprising an amino acid sequence beingat least 70%, optionally at least about 80%, preferably at least about85%, more preferably at least about 90% and most preferably at leastabout 95, 96, 97, 98 or 99% homologous to the sequenceFMLAAPSQREEEKKIWQGPGLLLCPHCNPHYHQY (SEQ ID NO: 292) of H68654_(—)1_P14(SEQ ID NO:40).

2. Comparison report between H68654_(—)1_P14 (SEQ ID NO:40) and knownproteins NP_(—)001034461 (SEQ ID NO: 35) (FIG. 30H):

A. An isolated chimeric polypeptide encoding for H68654_(—)1_P14 (SEQ IDNO:40), comprising a first amino acid sequence being at least 70%,optionally at least 80%, preferably at least 85%, more preferably atleast 90% and most preferably at least 95, 96, 97, 98 or 99%, homologousto a polypeptide having the sequence MWLKVFTTFLSFAT (SEQ ID NO: 289)corresponding to amino acids 1-14 of H68654_(—)1_P14 (SEQ ID NO:40), asecond amino acid sequence being at least 90% homologous toGACSGLKVTVPSHTVHGVRGQALYLPVHYGFHTPASDIQIIWLFERPHTMPKYLLGSVNKSVVPDLEYQHKFTMMPPNASLLINPLQFPDEGNYIVKVNIQGNGTLSASQKIQVTVDDPVTKPVVQIHPPSGAVEYVGNMTLTCHVEGGTRLAYQWLKNGRPVHTSSTYSFSPQNNTLHIAPVTKEDIGNYSCLVRNPVSEMESDIIMPIIYYGPYGLQVNSDKGLKVGEVFTVDLGEAILFDCSADSHPPNTYSWIRRTDNTTYIIKHGPRLEVASEKVAQKTMDYVCCAYNNITGRQDETHFTVIITSVGLEKLAQKGKSLSPLASITGISLFLIISMCLLFLWKKYQPYKVIKQKLEGRPETEYRKAQTFSG corresponding to aminoacids 27-401 of known proteins NP_(—)001034461 (SEQ ID NO: 35), whichalso corresponds to amino acids 15-389 of H68654_(—)1_P14 (SEQ IDNO:40), and a third amino acid sequence being at least 70%, optionallyat least 80%, preferably at least 85%, more preferably at least 90% andmost preferably at least 95, 96, 97, 98 or 99% homologous to apolypeptide having the sequence FMLAAPSQREEEKKIWQGPGLLLCPHCNPHYHQY (SEQID NO: 292) corresponding to amino acids 390-423 of H68654_(—)1_P14 (SEQID NO:40), wherein said first amino acid sequence, second amino acidsequence and third amino acid sequence are contiguous and in asequential order.

B. An isolated polypeptide encoding for a head of H68654_(—)1_P14 (SEQID NO:40), comprising a polypeptide being at least 70%, optionally atleast about 80%, preferably at least about 85%, more preferably at leastabout 90% and most preferably at least about 95, 96, 97, 98 or 99%homologous to the sequence MWLKVFTTFLSFAT (SEQ ID NO: 289) ofH68654_(—)1_P14 (SEQ ID NO:40).

C. An isolated polypeptide encoding for an edge portion ofH68654_(—)1_P14 (SEQ ID NO:40), comprising an amino acid sequence beingat least 70%, optionally at least about 80%, preferably at least about85%, more preferably at least about 90% and most preferably at leastabout 95, 96, 97, 98 or 99% homologous to the sequenceFMLAAPSQREEEKKIWQGPGLLLCPHCNPHYHQY (SEQ ID NO: 292) of H68654_(—)1_P14(SEQ ID NO:40).

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein H68654_(—)1_P14 (SEQ ID NO:40) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 85,(given according to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:40)).

TABLE 85 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 21 K −> R 74 K −> N 192 L −> P

The variant protein has the following domains, as determined by usingInterPro. The domains are described in Table 86:

TABLE 86 InterPro domains Analysis Domain description type Positions onprotein Immunoglobulin C2 type HMMSmart 152-214 Immunoglobulin subtypeHMMSmart 26-129, 143-223, 243-321 Immunoglobulin-like ProfileScan137-221, 223-319 Immunoglobulin V-set HMMPfam  19-131Immunoglobulin-like HMMPfam 151-209

Variant protein H68654_(—)1_P14 (SEQ ID NO:40) is encoded by thetranscript H68654_(—)1_T20 (SEQ ID NO:34), for which the coding portionstarts at position 102 and ends at position 1370. The transcript alsohas the following SNPs as listed in Table 87 (given according to theirposition on the nucleotide sequence, with the alternative nucleic acidlisted).

TABLE 87 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence A −> G 163 G −> A 323 G −> C 323 T −> C 551, 676 C −> A 1491  C−> G 1491 

As noted above, cluster H68654 features 3 segments, which were listed inTable 63. These segments are portions of nucleic acid sequences whichare described herein separately because they are of particular interest.A description of each segment according to the present invention is nowprovided.

Segment cluster H68654_(—)1_N3 (SEQ ID NO:123) according to the presentinvention is supported by 24 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: H68654_(—)1_TO (SEQ ID NO:25), H68654_(—)1_T15(SEQ ID NO:29), H68654_(—)1_T16 (SEQ ID NO:30), H68654_(—)1_T17 (SEQ IDNO:31), H68654_(—)1_T18 (SEQ ID NO:32), H68654_(—)1_T19 (SEQ ID NO:33),H68654_(—)1_T20 (SEQ ID NO:34), H68654_(—)1_T4 (SEQ ID NO:26),H68654_(—)1_T5 (SEQ ID NO:27) and H68654_(—)1_T8 (SEQ ID NO:28). Table88 below describes the starting and ending position of this segment oneach transcript.

TABLE 88 Segment location on transcripts Segment Segment Transcript namestarting position ending position H68654_1_T0 (SEQ ID NO: 25) 158 508H68654_1_T15 (SEQ ID NO: 29) 145 495 H68654_1_T16 (SEQ ID NO: 30) 145495 H68654_1_T17 (SEQ ID NO: 31) 145 495 H68654_1_T18 (SEQ ID NO: 32)145 495 H68654_1_T19 (SEQ ID NO: 33) 145 495 H68654_1_T20 (SEQ ID NO:34) 145 495 H68654_1_T4 (SEQ ID NO: 26) 145 495 H68654_1_T5 (SEQ ID NO:27) 158 508 H68654_1_T8 (SEQ ID NO: 28) 145 495

Segment cluster H68654_(—)1_N7 (SEQ ID NO:124) according to the presentinvention is supported by 20 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: H68654_(—)1_T0 (SEQ ID NO:25), H68654_(—)1_T15(SEQ ID NO:29), H68654_(—)1_T16 (SEQ ID NO:30), H68654_(—)1_T17 (SEQ IDNO:31), H68654_(—)1_T18 (SEQ ID NO:32), H68654_(—)1_T19 (SEQ ID NO:33),H68654_(—)1_T20 (SEQ ID NO:34), H68654_(—)1_T4 (SEQ ID NO:26),H68654_(—)1_T5 (SEQ ID NO:27) and H68654_(—)1_T8 (SEQ ID NO:28). Table89 below describes the starting and ending position of this segment oneach transcript.

TABLE 89 Segment location on transcripts Segment Segment Transcript namestarting position ending position H68654_1_T0 (SEQ ID NO: 25) 794 1090H68654_1_T15 (SEQ ID NO: 29) 781 1077 H68654_1_T16 (SEQ ID NO: 30) 7811077 H68654_1_T17 (SEQ ID NO: 31) 781 1077 H68654_1_T18 (SEQ ID NO: 32)781 1077 H68654_1_T19 (SEQ ID NO: 33) 781 1077 H68654_1_T20 (SEQ ID NO:34) 781 1077 H68654_1_T4 (SEQ ID NO: 26) 781 1077 H68654_1_T5 (SEQ IDNO: 27) 794 1090 H68654_1_T8 (SEQ ID NO: 28) 781 1077

Segment cluster H68654_(—)1_N12 (SEQ ID NO:125) according to the presentinvention is supported by 18 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: H68654_(—)1_T0 (SEQ ID NO:25), H68654_(—)1_T15(SEQ ID NO:29), H68654_(—)1_T16 (SEQ ID NO:30), H68654_(—)1_T17 (SEQ IDNO:31), H68654_(—)1_T18 (SEQ ID NO:32), H68654_(—)1_T19 (SEQ ID NO:33),H68654_(—)1_T20 (SEQ ID NO:34), H68654_(—)1_T4 (SEQ ID NO:26),H68654_(—)1_T5 (SEQ ID NO:27) and H68654_(—)1_T8 (SEQ ID NO:28). Table90 below describes the starting and ending position of this segment oneach transcript.

TABLE 90 Segment location on transcripts Segment Segment Transcript namestarting position ending position H68654_1_T0 (SEQ ID NO: 25) 1091 1216H68654_1_T15 (SEQ ID NO: 29) 1078 1203 H68654_1_T16 (SEQ ID NO: 30) 10781203 H68654_1_T17 (SEQ ID NO: 31) 1078 1203 H68654_1_T18 (SEQ ID NO: 32)1078 1203 H68654_1_T19 (SEQ ID NO: 33) 1078 1203 H68654_1_T20 (SEQ IDNO: 34) 1078 1203 H68654_1_T4 (SEQ ID NO: 26) 1078 1203 H68654_1_T5 (SEQID NO: 27) 1091 1216 H68654_1_T8 (SEQ ID NO: 28) 1078 1203

Expression of hypothetical protein LOC253012 H68654 transcripts whichare detectable by amplicon as depicted in sequence nameH68654_seg3WTF2R2 (SEQ ID NO: 226) in normal and cancerous Lung tissuesand in different normal tissues

Expression of hypothetical protein LOC253012 transcripts detectable byor according to seg3F2R2_ H68654_seg3WTF2R2 (SEQ ID NO: 226) ampliconand primers H68654_seg3WTF2 (SEQ ID NO: 224) and H68654_seg3WTR2 (SEQ IDNO: 225) was measured by real time PCR on lung panel and normal panel.The samples used are detailed in Table 3 and Table 2 above,respectively. For each RT sample, the expression of the above ampliconwas normalized to the normalization factor calculated from theexpression of several house keeping genes as described in Example 1.

Lung panel—The normalized quantity of each RT sample was then divided bythe median of the quantities of the normal samples (sample numbers 51-64and 69-70, Table 3 above), to obtain a value of fold up-regulation foreach sample relative to median of the normal samples.

FIG. 31 is a histogram showing over expression of the above-indicatedhypothetical protein LOC253012 transcripts in cancerous Lung samplesrelative to the normal samples.

As is evident from FIG. 31, the expression of hypothetical proteinLOC253012 transcripts detectable by the above amplicon in small cellcarcinoma samples was significantly higher than in the non-canceroussamples (sample numbers 51-64 and 69-70, Table 3 above). Notably anover-expression of at least 80 fold was found in 7 out of 9 small cellcarcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below.

The P value for the difference in the expression levels of hypotheticalprotein LOC253012 transcripts detectable by the above amplicon in Lungsmall cell carcinoma samples versus the normal tissue samples wasdetermined by T test as 3.24e-003.

Threshold of 80 fold over expression was found to differentiate betweensmall cell carcinoma and normal samples with P value of 7.49e-005 aschecked by exact Fisher test.

The above values demonstrate statistical significance of the results.

Normal panel—The normalized quantity of each RT sample was then dividedby the median of the quantities of the lung samples (sample numbers 26,28, 29 and 30, Table 2 above), to obtain a value of relative expressionof each sample relative to median of the lung samples, as shown in FIG.32.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: H68654_seg3WTF2 (SEQ ID NO: 224) forward primer;and H68654_seg3WTR2 (SEQ ID NO: 225) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: H68654_seg3WTF2R2 (SEQID NO: 226).

Forward Primer >H68654_seg3WTF2 (SEQ ID NO: 224): ATCACACACTGTCCATGGCGTReverse Primer >H68654_seg3WTR2 (SEQ ID NO: 225):GTCTCTCAAATAGCCATATGATCTGG Amplicon >H68654_seg3WTF2R2 (SEQ ID NO: 226)ATCACACACTGTCCATGGCGTCAGAGGTCAGGCCCTCTACCTACCCGTCCACTATGGCTTCCACACTCCAGCATCAGACATCCAGATCATATGGCTAT TTGAGAGAC

Expression of hypothetical protein LOC253012 H68654 transcripts whichare detectable by amplicon as depicted in sequence name H68654_seg7-12WT(SEQ ID NO: 223) in normal and cancerous Lung tissues and in differentnormal tissues

Expression of hypothetical protein LOC253012 transcripts detectable byor according to seg7-12—H68654_seg7-12WT (SEQ ID NO: 223) amplicon andprimers H68654_seg7-12WTF1 (SEQ ID NO: 221) and H68654_seg7-12WTR1 (SEQID NO: 222) was measured by real time PCR on lung panel and normalpanel. The samples used are detailed in Table 3 and Table 2 above,respectively. For each RT sample, the expression of the above ampliconwas normalized to the normalization factor calculated from theexpression of several house keeping genes as described in Example 1.

Lung panel—Non-detected samples (samples no. 30, 41, 78 and 92, Table 3)were assigned Ct value of 41 and were calculated accordingly. Thenormalized quantity of each RT sample was then divided by the median ofthe quantities of the normal samples (sample numbers 51-64 and 69-70,Table 3 above), to obtain a value of fold up-regulation for each samplerelative to median of the normal samples.

FIG. 33 is a histogram showing over expression of the above-indicatedhypothetical protein LOC253012 transcripts in cancerous Lung samplesrelative to the normal samples.

As is evident from FIG. 33, the expression of hypothetical proteinLOC253012 transcripts detectable by the above amplicon in small cellcarcinoma samples was significantly higher than in the non-canceroussamples (sample numbers 51-64 and 69-70, Table 3 above) and was higherin a few squamous cell carcinoma samples. Notably an over-expression ofat least 25 fold was found in 6 out of 9 small cell carcinoma samplesand in 4 out of 24 squamous cell carcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. The P value for the difference in theexpression levels of hypothetical protein LOC253012 transcriptsdetectable by the above amplicon in Lung small cell carcinoma samplesand Lung squamous cell carcinoma samples versus the normal tissuesamples was determined by T test as 1.24e-002 and 2.97e-002,respectively.

Threshold of 25 fold over expression was found to differentiate betweensmall cell carcinoma and normal samples with P value of 4.74e-004 aschecked by exact Fisher test.

The above values demonstrate statistical significance of the results.

Normal panel—Non-detected sample (sample no. 16) was assigned Ct valueof 41 and was calculated accordingly. The normalized quantity of each RTsample was then divided by the median of the quantities of the lungsamples (sample numbers 26, 28, 29 and 30, Table 2 above), to obtain avalue of relative expression of each sample relative to median of thelung samples, as shown in FIG. 34.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: H68654_seg7-12WTF1 (SEQ ID NO: 221) forwardprimer; and H68654_seg7-12WTR1 (SEQ ID NO: 222) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: H68654_seg7-12WT (SEQID NO: 223).

Forward Primer >H68654_seg7-12WTF1 (SEQ ID NO: 221):ATGGGCCTCGCTTAGAAGTTG Reverse Primer >H68654_seg7-12WTR1(SEQ ID NO: 222): TTCTGTGCAAGCTTCTCCAGTCAmplicon >H68654_seg7-12WT (SEQ ID NO: 223):ATGGGCCTCGCTTAGAAGTTGCATCTGAGAAAGTAGCCCAGAAGACAATGGACTATGTGTGCTGTGCTTACAACAACATAACCGGCAGGCAAGATGAAACTCATTTCACAGTTATCATCACTTCCGTAGGACTGGAGAAGCTTGCAC AGAA

Expression of LOC253012 H68654 transcripts which are detectable byamplicon as depicted in sequence name H68654_seg3WTF2R2 (SEQ ID NO: 226)in the blood-specific panel.

Expression of LOC253012 transcripts detectable by or according toseg3-H68654seg3F2R2 (SEQ ID NO: 226) amplicon and primers H68654seg3F2(SEQ ID NO: 224) and H68654seg3R2 (SEQ ID NO: 225) was measured by realtime PCR on blood panel. The samples used are detailed in Table 1 above.For each RT sample, the expression of the above amplicon was normalizedto the normalization factor calculated from the expression of severalhouse keeping genes as described in Example 1. The normalized quantityof each RT sample was then divided by the median of the quantities ofthe kidney normal samples (sample numbers 65-67, Table 1 above), toobtain a value of relative expression of each sample relative to medianof the kidney normal samples.

The results of this analysis are depicted in the histogram in FIG. 35A.Expression of the above-indicated LOC253012 transcript is high in thekidney normal, colon normal and small intestine normal as in few of thedifferent blood samples checked.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: H68654_seg3WTF2 forward primer; andH68654_seg3WTR2 reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: H68654_seg3WTF2R2.

Forward Primer >H68654_seg3WTF2: (SEQ ID NO: 224) ATCACACACTGTCCATGGCGTReverse Primer >H68654_seg3WTR2: (SEQ ID NO: 225)GTCTCTCAAATAGCCATATGATCTGG  Amplicon >H68654_seg3WTF2R2 (SEQ ID NO: 226)ATCACACACTGTCCATGGCGTCAGAGGTCAGGCCCTCTACCTACCCGTCCACTATGGCTTCCACACTCCAGCATCAGACATCCAGATCATATGGCT ATTTGAGAGAC

Expression of LOC253012 H68654 transcripts which are detectable byamplicon as depicted in sequence name H68654seg7-12 (SEQ ID NO: 223) inthe blood-specific panel.

Expression of LOC253012 transcripts detectable by or according toseg7-12-H68654seg7-12WTF1R1 (SEQ ID NO: 223) amplicon and primersH68654seg7-12WTF1 (SEQ ID NO: 221) and H68654seg7-12WTR1 (SEQ ID NO:222) was measured by real time PCR on blood panel. The samples used aredetailed in Table 1 above. For each RT sample, the expression of theabove amplicon was normalized to the normalization factor calculatedfrom the expression of several house keeping genes as described inExample 1. The normalized quantity of each RT sample was then divided bythe median of the quantities of the kidney normal samples (samplenumbers 65-67, Table 1 above), to obtain a value of relative expressionof each sample relative to median of the kidney normal samples.

The results of this analysis are depicted in the histogram in FIG. 35B.Expression of the above-indicated LOC253012 transcript is higher in thekidney normal, colon normal and small intestine normal relative to thedifferent blood samples checked.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: seg7-12WTF1 forward primer (SEQ ID NO: 221); andseg7-12WTR1 reverse primer (SEQ ID NO: 222).

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: seg7-12WTF1R1 (SEQ IDNO: 223).

Forward Primer >H68654_seg7-12WTF1 (SEQ ID NO: 221)ATGGGCCTCGCTTAGAAGTTG Reverse Primer >H68654_seg7-12WTR1(SEQ ID NO: 222) TTCTGTGCAAGCTTCTCCAGTC Amplicon >H68654_seg7-12WT(SEQ ID NO: 223) ATGGGCCTCGCTTAGAAGTTGCATCTGAGAAAGTAGCCCAGAAGACAATGGACTATGTGTGCTGTGCTTACAACAACATAACCGGCAGGCAAGATGAAACTCATTTCACAGTTATCATCACTTCCGTAGGACTGGAGAAGCTTGCAC AGAA

Expression of hypothetical protein LOC253012 H68654 transcripts whichare detectable by amplicon as depicted in sequence name H68654_seg0-3(SEQ ID NO: 229) in normal and cancerous Lung tissues Expression ofLOC253012 transcripts detectable by or according to seg0-3-H68654_seg0-3(SEQ ID NO: 229) amplicon and primers H68654_seg0-3F1 (SEQ ID NO: 227)and H68654_seg0-3R1 (SEQ ID NO: 228) was measured by real time PCR onlung panel. The samples used are detailed in Table 3 above. For each RTsample, the expression of the above amplicon was normalized to thenormalization factor calculated from the expression of several housekeeping genes as described in Example 1. The normalized quantity of eachRT sample was then divided by the median of the quantities of the normalsamples (sample numbers 51-54, 56-64, 69 and 70, Table 3 above), toobtain a value of fold up-regulation for each sample relative to medianof the normal samples.

FIG. 36 is a histogram showing over expression of the above-indicatedLOC253012 transcripts in cancerous Lung samples relative to the normalsamples.

As is evident from FIG. 36, the expression of LOC253012 transcriptsdetectable by the above amplicon in squamous cell carcinoma and smallcell carcinoma samples was significantly higher than in thenon-cancerous samples (sample numbers 51-54, 56-64, 69 and 70, Table 3above). Notably an over-expression of at least 5 fold was found in 6 outof 21 squamous cell carcinoma samples and in 8 out of 9 small cellcarcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. The P value for the difference in theexpression levels of LOC253012 transcripts detectable by the aboveamplicon in lung squamous cell carcinoma samples and lung small cellcarcinoma samples versus the normal tissue samples was determined by Ttest as 4.96e-002 and 1.05e-003, respectively.

Threshold of 5 fold over expression was found to differentiate betweensquamous cell carcinoma and small cell carcinoma and normal samples withP value of 2.79e-002 and 1.22e-005, respectively, as checked by exactFisher test. The above values demonstrate statistical significance ofthe results.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: H68654_seg0-3F1 (SEQ ID NO: 227) forward primer;and H68654_seg0-3R1 (SEQ ID NO: 228) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: H68654_seg0-3F1R1 (SEQID NO: 229).

Forward Primer >H68654_seg0-3F1 (SEQ ID NO: 227) GCTTTCATGGAGCCCTTCGReverse Primer >H68654_seg0-3R1 (SEQ ID NO: 228) GCCTGACCTCTGACGCCAAmplicon >H68654_seg0-3F1R1 (SEQ ID NO: 229)GCTTTCATGGAGCCCTTCGGTGACACACTTGGGGTCTTTCAGTGCAAAATATACCTCCTTCTCTTCGGTGCTTGCTCGGGGCTGAAGGTGACAGTGCCATCACACACTGTCCATGGCGTCAGAGGTCAGGC

Expression of hypothetical protein LOC253012 H68654 transcripts whichare detectable by amplicon as depicted in sequence name H68654_seg2-3(SEQ ID NO: 232) in normal and cancerous Lung tissues

Expression of LOC253012 transcripts detectable by or according toseg2-3-H68654_seg2-3 (SEQ ID NO: 232) amplicon and primersH68654_seg2-3F1 (SEQ ID NO: 230) and H68654_seg2-3R1 (SEQ ID NO: 231)was measured by real time PCR on lung panel. The samples used aredetailed in Table 3 above. For each RT sample, the expression of theabove amplicon was normalized to the normalization factor calculatedfrom the expression of several house keeping genes as described inExample 1. The normalized quantity of each RT sample was then divided bythe median of the quantities of the normal samples (sample numbers 51,52, 54-64, 69 and 70, Table 3 above), to obtain a value of foldup-regulation for each sample relative to median of the normal samples.

FIG. 37 is a histogram showing over expression of the above-indicatedLOC253012 transcripts in cancerous Lung samples relative to the normalsamples.

As is evident from FIG. 37, the expression of LOC253012 transcriptsdetectable by the above amplicon in small cell carcinoma samples wassignificantly higher than in the non-cancerous samples (sample numbers51, 52, 54-64, 69 and 70, Table 3 above) Notably an over-expression ofat least 8 fold was found in 7 out of 9 small cell carcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. The P value for the difference in theexpression levels of LOC253012 transcripts detectable by the aboveamplicon in Lung small cell carcinoma samples versus the normal tissuesamples was determined by T test as 1.93e-002.

Threshold of 8 fold over expression was found to differentiate betweensmall cell carcinoma and normal samples with P value of 1.04e-004 aschecked by exact Fisher test.

The above values demonstrate statistical significance of the results.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: H68654_seg2-3F1 (SEQ ID NO: 230) forward primer;and H68654_seg2-3R1 (SEQ ID NO: 231) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: H68654_seg2-3F1R1 (SEQID NO: 232).

Forward Primer >H68654_seg2-3F1 (SEQ ID NO: 230) CTCTGCATTTGCCCCTTTAGAReverse Primer >H68654_seg2-3R1 (SEQ ID NO: 231) GATGGCACTGTCACCTTCAGCAmplicon >H68654_seg2-3F1R1 (SEQ ID NO: 232)CTCTGCATTTGCCCCTTTAGATTGTGAAATGTGGCTCAAGGTCTTCACAACTTTCCTTTCCTTTGCAACAGGTGCTTGCTCGGGGCTGAAGGTGACAG TGCCATC

Example 6 Description for Cluster H19011_(—)1

The present invention relates to C1ORF32 polypeptides, novel splicevariants and diagnostics and therapeutics based thereon.

Cluster H19011_(—)1 (internal ID 76432827) features 2 transcripts and 5segments of interest, the names for which are given in Tables 91 and 92,respectively. The selected protein variants are given in table 93.

TABLE 91 Transcripts of interest Transcript Name H19011_1_T8 (SEQ ID NO:45) H19011_1_T9 (SEQ ID NO: 46)

TABLE 92 Segments of interest Segment Name H19011_1_N13 (SEQ ID NO: 129)H19011_1_N8 (SEQ ID NO: 130) H19011_1_N10 (SEQ ID NO: 131) H19011_1_N11(SEQ ID NO: 132) H19011_1_N12 (SEQ ID NO: 133)

TABLE 93 Proteins of interest Protein Name Corresponding TranscriptsH19011_1_P8 (SEQ ID NO: 48) H19011_1_T8 (SEQ ID NO: 45) H19011_1_P9 (SEQID NO: 50) H19011_1_T9 (SEQ ID NO: 46)

These sequences are variants of the known protein hypothetical proteinLOC387597 (RefSeq accession identifier NP_(—)955383 (SEQ ID NO: 47),synonyms: C1ORF32, NP_(—)955383; LISCH-like; RP4-782G3.2; dJ782G3.1),referred to herein as the previously known protein.

C1ORF32 is a hypothetical protein that was computationally discoveredduring the annotation of chromosome 1 (Gregory S G et al. 2006, Nature441 (7091) 315-321). Its closest annotated homolog belongs to the LISCH7family, a subfamily of the immunoglobulin super family. One of theannotated members of this family is the lipolysis-stimulated lipoproteinreceptor which has a probable role in the clearance of triglyceride-richlipoprotein from blood (Swissprot annotation of accession Q86X29).

According to the present invention, C1ORF32 was predicted to be a novelB7/CD28 member based on the presence of an IgV domain, in addition ofits being a type I membrane protein, like other known B7 members. Also,two alternatively spliced variants of the present invention(H19011_(—)1_P8 (SEQ ID NO:48) and H19011_(—)1_P9 (SEQ ID NO:50)), whichshare only the first 5 exons with the wild type C1ORF32, are similar tothe known B7 family members in their exons' sizes and the position ofthe IgV and transmembrane domains within these exons. In addition,C1ORF32 was shown in the present invention to be overexpressed in smallcell lung cancer.

As noted above, cluster H19011 features 2 transcripts, which were listedin Table 91 above. These transcripts encode for proteins which arevariants of protein hypothetical protein LOC387597 (SEQ ID NO:47). Adescription of each variant protein according to the present inventionis now provided.

Variant protein H19011_(—)1_P8 (SEQ ID NO:48) according to the presentinvention has an amino acid sequence as encoded by transcriptH19011_(—)1_T8 (SEQ ID NO:45). Alignments to one or more previouslypublished protein sequences are shown in FIG. 38A. A brief descriptionof the relationship of the variant protein according to the presentinvention to each such aligned protein is as follows:

Comparison report between H19011_(—)1_P8 (SEQ ID NO:48) and knownproteins Q71H61_HUMAN and NP_(—)955383 (SEQ ID NO: 47) (FIG. 38A):

A. An isolated chimeric polypeptide encoding for H19011_(—)1_P8 (SEQ IDNO:48), comprising a first amino acid sequence being at least 90%homologous to MDRVLLRWISLFWLTAMVEGLQVTVPDKKKVAMLFQPTVLRCHFSTSSHQPAVVQWKFKSYCQDRMGESLGMSSTRAQSLSKRNLEWDPYLDCLDSRRTVRVVASKQGSTVTLGDFYRGREITIVHDADLQIGKLMWGDSGLYYCIITTPDDLEGKNE corresponding toamino acids 1-158 of known proteins Q71H61_HUMAN and NP_(—)955383 (SEQID NO: 47), which also corresponds to amino acids 1-158 ofH19011_(—)1_P8 (SEQ ID NO:48), a bridging amino acid G corresponding toamino acid 159 of H19011_(—)1_P8 (SEQ ID NO:48), a second amino acidsequence being at least 90% homologous to S corresponding to amino acids160-160 of known proteins Q71H61_HUMAN and NP_(—)955383 (SEQ ID NO: 47),which also corresponds to amino acids 160-160 of H19011_(—)1_P8 (SEQ IDNO:48), bridging amino acids LG corresponding to amino acid 161-162 ofH19011_(—)1_P8 (SEQ ID NO:48), a third amino acid sequence being atleast 90% homologous toLLVLGRTGLLADLLPSFAVEIMPEWVFVGLVLLGVFLFFVLVGICWCQCCPHSCC CYVRCPCCPDSCcorresponding to amino acids 163-229 of known proteins Q71H61_HUMAN andNP_(—)955383 (SEQ ID NO: 47), which also corresponds to amino acids163-229 of H19011_(—)1_P8 (SEQ ID NO:48), a bridging amino acid Wcorresponding to amino acid 230 of H19011_(—)1_P8 (SEQ ID NO:48), afourth amino acid sequence being at least 90% homologous to CPQAcorresponding to amino acids 231-234 of known proteins Q71H61_HUMAN andNP_(—)955383 (SEQ ID NO: 47), which also corresponds to amino acids231-234 of H19011_(—)1_P8 (SEQ ID NO:48), and a fifth amino acidsequence being at least 70%, optionally at least 80%, preferably atleast 85%, more preferably at least 90% and most preferably at least 95,96, 97, 98 or 99% homologous to a polypeptide having the sequenceCEYSDRWGDRAIERNVYLST (SEQ ID NO: 293) corresponding to amino acids235-254 of H19011_(—)1_P8 (SEQ ID NO:48), wherein said first amino acidsequence, bridging amino acid, second amino acid sequence, bridgingamino acid, third amino acid sequence, bridging amino acid, fourth aminoacid sequence and fifth amino acid sequence are contiguous and in asequential order.

B. An isolated polypeptide encoding for an edge portion ofH19011_(—)1_P8 (SEQ ID NO:48), comprising an amino acid sequence beingat least 70%, optionally at least about 80%, preferably at least about85%, more preferably at least about 90% and most preferably at leastabout 95, 96, 97, 98 or 99% homologous to the sequenceCEYSDRWGDRAIERNVYLST (SEQ ID NO: 293) of H19011_(—)1_P8 (SEQ ID NO:48).

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein H19011_(—)1_P8 (SEQ ID NO:48) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 94,(given according to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:48)). An example of such adeduced sequence, with alternative amino-acids, that was produced (usingpart of the SNPs below), is given under the name H19011_(—)1_P8_V1 (SEQID NO:49).

TABLE 94 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 159 G −> D 161 L −> V 162 G −> E 202 V −> D 202 V −>G 230 W −> C

The variant protein has the following domains, as determined by usingInterPro. The domains are described in Table 95:

TABLE 95 InterPro domains Analysis Domain description type Positions onprotein LISCH7 HMMPfam 186-234 IG SMART  27-166

Variant protein H19011_(—)1_P8 (SEQ ID NO:48) is encoded by thetranscript H19011_(—)1_T8 (SEQ ID NO:45), for which the coding portionstarts at position 181 and ends at position 942. The transcript also hasthe following SNPs as listed in Table 96 (given according to theirposition on the nucleotide sequence, with the alternative nucleic acidlisted).

TABLE 96 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence G −> A 656 C −> G 661 G −> A 665 T −> A 785 T −> G 785 G −> C870

The genomic structure of protein H19011_(—)1_P8 (SEQ ID NO:48) (numberof exons relevant to the extra-cellular region of the protein, thelength of these exons, the frame of the codon in which the introns areinserted and the location of the protein features and domains in thegene structure) is characteristic to the ligands of theB7/co-stimulatory protein family, as given in table 97

TABLE 97 genomic structure and protein features Exon Exon Amino- numberLength Acids Protein feature on exon 1 46  1-15 Signal Peptide 2 333 16-126 IgV domain 3 120 127-166 IgC2 domain 4 57 167-185 5 206 186-254Trans-membrane region

Variant protein H19011_(—)1_P9 (SEQ ID NO:50) according to the presentinvention has an amino acid sequence as encoded by transcriptH19011_(—)1_T9 (SEQ ID NO:46). Alignments to one or more previouslypublished protein sequences are shown in FIG. 38B. A brief descriptionof the relationship of the variant protein according to the presentinvention to each such aligned protein is as follows:

Comparison report between H19011_(—)1_P9 (SEQ ID NO:50) and knownproteins Q71H61_HUMAN and NP_(—)955383 (SEQ ID NO: 47) (FIG. 38B):

A. An isolated chimeric polypeptide encoding for H19011_(—)1_P9 (SEQ IDNO:50), comprising a first amino acid sequence being at least 90%homologous to MDRVLLRWISLFWLTAMVEGLQVTVPDKKKVAMLFQPTVLRCHFSTSSHQPAVVQWKFKSYCQDRMGESLGMSSTRAQSLSKRNLEWDPYLDCLDSRRTVRVVASKQGSTVTLGDFYRGREITIVHDADLQIGKLMWGDSGLYYCIITTPDDLEGKNE corresponding toamino acids 1-158 of known proteins Q71H61_HUMAN and NP_(—)955383 (SEQID NO: 47), which also corresponds to amino acids 1-158 ofH19011_(—)1_P9 (SEQ ID NO:50), a bridging amino acid G corresponding toamino acid 159 of H19011_(—)1_P9 (SEQ ID NO:50), a second amino acidsequence being at least 90% homologous to S corresponding to amino acids160-160 of known proteins Q71H61_HUMAN and NP_(—)955383 (SEQ ID NO: 47),which also corresponds to amino acids 160-160 of H19011_(—)1_P9 (SEQ IDNO:50), bridging amino acids LG corresponding to amino acid 161-162 ofH19011_(—)1_P9 (SEQ ID NO:50), a third amino acid sequence being atleast 90% homologous to LLVL corresponding to amino acids 163-166 ofknown proteins Q71H61_HUMAN and NP_(—)955383 (SEQ ID NO: 47), which alsocorresponds to amino acids 163-166 of H19011_(—)1_P9 (SEQ ID NO:50), afourth amino acid sequence being at least 90% homologous toEWVFVGLVLLGVFLFFVLVGICWCQCCPHSCCCYVRCPCCPDSC corresponding to aminoacids 186-229 of known proteins Q71H61_HUMAN and NP_(—)955383 (SEQ IDNO: 47), which also corresponds to amino acids 167-210 of H19011_(—)1_P9(SEQ ID NO:50), a bridging amino acid W corresponding to amino acid 211of H19011_(—)1_P9 (SEQ ID NO:50), a fifth amino acid sequence being atleast 90% homologous to CPQA corresponding to amino acids 231-234 ofknown proteins Q71H61_HUMAN and NP_(—)955383 (SEQ ID NO: 47), which alsocorresponds to amino acids 212-215 of H19011_(—)1_P9 (SEQ ID NO:50), anda sixth amino acid sequence being at least 70%, optionally at least 80%,preferably at least 85%, more preferably at least 90% and mostpreferably at least 95, 96, 97, 98 or 99% homologous to a polypeptidehaving the sequence CEYSDRWGDRAIERNVYLST (SEQ ID NO: 293) correspondingto amino acids 216-235 of H19011_(—)1_P9 (SEQ ID NO:50), wherein saidfirst amino acid sequence, bridging amino acid, second amino acidsequence, bridging amino acid, third amino acid sequence, fourth aminoacid sequence, bridging amino acid, fifth amino acid sequence and sixthamino acid sequence are contiguous and in a sequential order.

B. An isolated chimeric polypeptide encoding for an edge portion ofH19011_(—)1_P9 (SEQ ID NO:50), comprising a polypeptide having a length“n”, wherein n is at least about 10 amino acids in length, optionally atleast about 20 amino acids in length, preferably at least about 30 aminoacids in length, more preferably at least about 40 amino acids in lengthand most preferably at least about 50 amino acids in length, wherein atleast two amino acids comprise LE, having a structure as follows: asequence starting from any of amino acid numbers 166−x to 166; andending at any of amino acid numbers 167+((n−2)−x), in which x variesfrom 0 to n−2.

C. An isolated polypeptide encoding for an edge portion ofH19011_(—)1_P9 (SEQ ID NO:50), comprising an amino acid sequence beingat least 70%, optionally at least about 80%, preferably at least about85%, more preferably at least about 90% and most preferably at leastabout 95, 96, 97, 98 or 99% homologous to the sequenceCEYSDRWGDRAIERNVYLST (SEQ ID NO: 293) of H19011_(—)1_P9 (SEQ ID NO:50).

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein H19011_(—)1_P9 (SEQ ID NO:50) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table 98,(given according to their positions on the amino acid sequence, with thealternative amino acids listed (SEQ ID NO:50)).

TABLE 98 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 159 G −> D 161 L −> V 162 G −> E 183 V −> D 183 V −>G 211 W −> C

Variant protein H19011_(—)1_P9 (SEQ ID NO:50) is encoded by thetranscript H19011_(—)1_T9 (SEQ ID NO:46), for which the coding portionstarts at position 181 and ends at position 885. The transcript also hasthe following SNPs as listed in Table 99 (given according to theirposition on the nucleotide sequence, with the alternative nucleic acidlisted).

TABLE 99 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence G −> A 656 C −> G 661 G −> A 665 T −> A 728 T −> G 728 G −> C813

As noted above, cluster H19011 features 5 segments, which were listed inTable 92 above. These segments are portions of nucleic acid sequenceswhich are described herein separately because they are of particularinterest. A description of each segment according to the presentinvention is now provided.

Segment cluster H19011_(—)1_N13 (SEQ ID NO:129) according to the presentinvention is supported by 3 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: H19011_(—)1_T8 (SEQ ID NO:45) and H19011_(—)1_T9(SEQ ID NO:46). Table 100 below describes the starting and endingposition of this segment on each transcript.

TABLE 100 Segment location on transcripts Segment Segment Transcriptname starting position ending position H19011_1_T8 (SEQ ID NO: 45) 8841407 H19011_1_T9 (SEQ ID NO: 46) 827 1350

According to an optional embodiment of the present invention, shortsegments related to the above cluster are also provided. These segmentsare up to about 120 bp in length, and so are included in a separatedescription.

Segment cluster H19011_(—)1_N8 (SEQ ID NO:130) according to the presentinvention is supported by 4 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: H19011_(—)1_T8 (SEQ ID NO:45). Table 101 belowdescribes the starting and ending position of this segment on eachtranscript.

TABLE 101 Segment location on transcripts Segment Segment Transcriptname starting position ending position H19011_1_T8 (SEQ ID NO: 45) 680736

Segment cluster H19011_(—)1_N10 (SEQ ID NO:131) according to the presentinvention is supported by 5 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: H19011_(—)1_T8 (SEQ ID NO:45) and H19011_(—)1_T9(SEQ ID NO:46). Table 102 below describes the starting and endingposition of this segment on each transcript.

TABLE 102 Segment location on transcripts Segment Segment Transcriptname starting position ending position H19011_1_T8 (SEQ ID NO: 45) 737797 H19011_1_T9 (SEQ ID NO: 46) 680 740

Segment cluster H19011_(—)1_N11 (SEQ ID NO:132) according to the presentinvention is supported by 3 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: H19011_(—)1_T8 (SEQ ID NO:45) and H19011_(—)1_T9(SEQ ID NO:46). Table 103 below describes the starting and endingposition of this segment on each transcript.

TABLE 103 Segment location on transcripts Segment Segment Transcriptname starting position ending position H19011_1_T8 (SEQ ID NO: 45) 798863 H19011_1_T9 (SEQ ID NO: 46) 741 806

Segment cluster H19011_(—)1_N12 (SEQ ID NO:133) according to the presentinvention is supported by 5 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: H19011_(—)1_T8 (SEQ ID NO:45) and H19011_(—)1_T9(SEQ ID NO:46). Table 104 below describes the starting and endingposition of this segment on each transcript.

TABLE 104 Segment location on transcripts Segment Segment Transcriptname starting position ending position H19011_1_T8 (SEQ ID NO: 45) 864883 H19011_1_T9 (SEQ ID NO: 46) 807 826

Expression of C1ORF32, chromosome 1 open reading frame 32, H19011transcripts which are detectable by amplicon as depicted in sequencename H19011_seg13F2R2 (SEQ ID NO: 235) in normal and cancerous Colontissues, in normal and cancerous Lung tissues and in different normaltissues

Expression of C1ORF32, chromosome 1 open reading frame 32, transcriptsdetectable by or according to seg13—H19011_seg13F2R2 (SEQ ID NO: 235)amplicon and primers H19011_seg13F2 (SEQ ID NO: 233) and H19011_seg13R2(SEQ ID NO: 234) was measured by real time PCR on colon panel, lungpanel and normal panel. The samples used are detailed in Table 5, Table3 and Table 2 above, respectively. For each RT sample, the expression ofthe above amplicon was normalized to the normalization factor calculatedfrom the expression of several house keeping genes as described inExample 1.

Colon panel—The normalized quantity of each RT sample was then dividedby the median of the quantities of the normal samples (sample numbers42-70, Table 5 above). Then the reciprocal of this ratio was calculated,to obtain a value of fold down-regulation for each sample relative tomedian of the normal samples.

FIG. 39 is a histogram showing down regulation of the above-indicatedC1ORF32 transcripts in cancerous Colon samples relative to the normalsamples.

As is evident from FIG. 39, the expression of C1ORF32 transcriptsdetectable by the above amplicon in cancer samples was significantlylower than in the non-cancerous samples (sample numbers 42-70, Table 5above). Notably down regulation of at least 6 fold was found in 17 outof 55 adenocarcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. The P value for the difference in theexpression levels of C1ORF32 transcripts detectable by the aboveamplicon in Colon cancer samples versus the normal tissue samples wasdetermined by T test as 9.36e-004.

Threshold of 6 fold down regulation was found to differentiate betweencancer and normal samples with P value of 2.67e-004 as checked by exactFisher test.

The above values demonstrate statistical significance of the results.

Lung panel—The normalized quantity of each RT sample was then divided bythe median of the quantities of the normal samples (sample numbers 51-64and 69-70, Table 3 above), to obtain a value of fold up-regulation foreach sample relative to median of the normal samples.

FIG. 40 is a histogram showing over expression of the above-indicatedC1ORF32 transcripts in cancerous Lung samples relative to the normalsamples.

As is evident from FIG. 40, the expression of C1ORF32 transcriptsdetectable by the above amplicon in small cell carcinoma samples wassignificantly higher than in the non-cancerous samples (sample numbers51-64 and 69-70, Table 3 above). Notably an over-expression of at least6 fold was found in 9 out of 9 small cell carcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below.

The P value for the difference in the expression levels of C1ORF32transcripts detectable by the above amplicon in Lung small cellcarcinoma samples versus the normal tissue samples was determined by Ttest as 3.43e-003.

Threshold of 6 fold over expression was found to differentiate betweensmall cell carcinoma and normal samples with P value of 4.89e-007 aschecked by exact Fisher test.

The above values demonstrate statistical significance of the results.

Normal panel—The normalized quantity of each RT sample was then dividedby the median of the quantities of the colon samples (sample numbers 3,4 and 5, Table 2 above), to obtain a value of relative expression ofeach sample relative to median of the colon samples, as shown in FIG.41A. The normalized quantity of each RT sample was then divided by themedian of the quantities of the lung samples (sample numbers 26, 28, 29and 30, Table 2 above), to obtain a value of relative expression of eachsample relative to median of the lung samples, as shown in FIG. 41B.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: H19011_seg13F2 (SEQ ID NO: 233) forward primer;and H19011_seg13R2 (SEQ ID NO: 234) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: H19011_seg13F2R2 (SEQID NO: 235).

Forward Primer >H19011_seg13F2 (SEQ ID NO: 233): GTGAGTACAGTGACCGCTGGGReverse Primer >H19011_seg13R2 (SEQ ID NO: 234): GGAGAAGAGTCTGGAATGACCAAAmplicon >H19011_seg13F2R2 (SEQ ID NO: 235)GTGAGTACAGTGACCGCTGGGGAGACAGAGCGATCGAGAGAAATGTCTACCTCTCTACCTGACAGCTGTGTGCGCTGGGTTCCTCCTCCACCTCCTGTCCTGCCACCCCCAAGATTGGTCATTCCAGACTCTTCTCC

Expression of C1ORF32, chromosome 1 open reading frame 32, H19011transcripts which are detectable by amplicon as depicted in sequencename H19011_seg8-13F1R1 (SEQ ID NO: 238) in normal and cancerous Lungtissues

Expression of C1ORF32, chromosome 1 open reading frame 32, transcriptsdetectable by or according to seg8-13F1R1_ H19011_seg8-13F1R1 (SEQ IDNO: 238) amplicon and primers H19011_seg8-13F1 (SEQ ID NO: 236) andH19011_seg8-13R1 (SEQ ID NO: 237) was measured by real time PCR on lungpanel. The samples used are detailed in Table 3 above. Samples thatshowed no detection of the amplicon (samples no. 1, 2, 4-10, 12-27,29-35, 37-41, 51-64 and 69-70, Table 3) were assigned Ct value of 41 andwere calculated accordingly. These samples showed a primer-dimer productwith a characteristic dissociation curve and a significantly lower TM(this artefactual product was identified by its appearance in thenegative control without RT sample). For each RT sample, the expressionof the above amplicon was normalized to the normalization factorcalculated from the expression of several house keeping genes asdescribed in Example 1. The normalized quantity of each RT sample wasthen divided by the median of the quantities of the normal samples(sample numbers 51-64 and 69-70, Table 3 above), to obtain a value offold up-regulation for each sample relative to median of the normalsamples.

FIG. 42 is a histogram showing over expression of the above-indicatedC1ORF32 transcripts in cancerous Lung samples relative to the normalsamples.

As is evident from FIG. 42, the expression of C1ORF32 transcriptsdetectable by the above amplicon in small cell carcinoma samples wassignificantly higher than in the non-cancerous samples (sample numbers51-64 and 69-70, Table 3 above). Notably an over-expression of at least500 fold was found in 9 out of 9 small cell carcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. The P value for the difference in theexpression levels of C1ORF32 transcripts detectable by the aboveamplicon in Lung small cell carcinoma samples versus the normal tissuesamples was determined by T test as 6.70e-003.

Threshold of 500 fold over expression was found to differentiate betweensmall cell carcinoma and normal samples with P value of 4.89e-007 aschecked by exact Fisher test.

The above values demonstrate statistical significance of the results.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: H19011_seg8-13F1 (SEQ ID NO: 236) forward primer;and H19011_seg8-13R1 (SEQ ID NO: 237) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: H19011_seg8-13F1R1(SEQ ID NO: 238).

Forward Primer >H19011_seg8-13F1 (SEQ ID NO: 236): GCCCAGTTTTGCTGTGGAGAReverse Primer >H19011_seg8-13R1 (SEQ ID NO: 237):GGTAGACATTTCTCTCGATCGCTC Amplicon >H19011_seg8-13F1R1 (SEQ ID NO: 238)GCCCAGTTTTGCTGTGGAGATTATGCCAGAGTGGGTGTTTGTTGGCCTGGTGCTCCTGGGCGTCTTCCTCTTCTTCGTCCTGGTGGGGATCTGCTGGTGCCAGTGCTGCCCTCACAGCTGCTGCTGCTATGTCCGCTGCCCATGCTGCCCAGATTCCTGCTGGTGCCCTCAAGCCTGTGAGTACAGTGACCGCTGGGGAGACAGAGCGATCGAGAGAAATGTCTACC

Expression of C1ORF32, chromosome 1 open reading frame 32, H19011transcripts which are detectable by amplicon as depicted in sequencename H19011_junc8-10seg13 (SEQ ID NO: 241) in normal and cancerous lungtissues, in normal and cancerous colon tissues, in different normaltissues and in the blood-specific panel.

Expression of C1ORF32 transcripts detectable by or according tojunc8-10seg13—H19011_junc8-10seg13 (SEQ ID NO: 241) amplicon and primersH19011_junc8-10seg13F1 (SEQ ID NO: 239) and H19011_junc8-10seg13R1 (SEQID NO: 240) was measured by real time PCR lung panel, colon panel,normal panel and blood panel. The samples used are detailed in Table 3,Table 5, Table 2 and Table 1 above, respectively. For each RT sample,the expression of the above amplicon was normalized to the normalizationfactor calculated from the expression of several house keeping genes asdescribed in Example 1.

For lung panel—Non-detected sample (sample no. 69, Table 3) was assignedCt value of 41 and was calculated accordingly. The normalized quantityof each RT sample was then divided by the median of the quantities ofthe normal samples (sample numbers 51, 53, 54, 56, 57, 59, 61, 62, 64and 70, Table 3 above), to obtain a value of fold up-regulation for eachsample relative to median of the normal samples.

FIG. 43 is a histogram showing over expression of the above-indicatedC1ORF32 transcripts in cancerous Lung samples relative to the normalsamples.

As is evident from FIG. 43, the expression of C1ORF32 transcriptsdetectable by the above amplicon in small cell carcinoma samples wassignificantly higher than in the non-cancerous samples (sample numbers51, 53, 54, 56, 57, 59, 61, 62, 64 and 70, Table 3 above). Notably anover-expression of at least 7 fold was found in 9 out of 9 small cellcarcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below.

The P value for the difference in the expression levels of C1ORF32transcripts detectable by the above amplicon in Lung small cellcarcinoma samples versus the normal tissue samples was determined by Ttest as 2.34e-003.

Threshold of 7 fold over expression was found to differentiate betweensmall cell carcinoma and normal samples with P value of 1.08e-005 aschecked by exact Fisher test.

The above values demonstrate statistical significance of the results.

For colon panel—Non-detected sample (sample no. 79, Table 5) wasassigned Ct value of 41 and was calculated accordingly. The normalizedquantity of each RT sample was then divided by the median of thequantities of the normal samples (sample numbers 42-62 and 65-70, Table5 above). Then the reciprocal of this ratio was calculated, to obtain avalue of fold down-regulation for each sample relative to median of thenormal samples.

FIG. 44 is a histogram showing down regulation of the above-indicatedC1ORF32 transcripts in cancerous colon samples relative to the normalsamples.

As is evident from FIG. 44, the expression of C1ORF32 transcriptsdetectable by the above amplicon in cancer samples was significantlylower than in the non-cancerous samples (sample numbers 42-62 and 65-70,Table 5 above). Notably down regulation of at least 5 fold was found in15 out of 36 adenocarcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. Threshold of 5 fold down regulation wasfound to differentiate between cancer and normal samples with P value of4.29e-004 as checked by exact Fisher test.

The above values demonstrate statistical significance of the results.

For normal panel—Non-detected samples (samples no. 42 and 49, Table 2)were assigned Ct value of 41 and were calculated accordingly. Thenormalized quantity of each RT sample was then divided by the median ofthe quantities of the colon samples (sample numbers 4 and 5, Table 2above), to obtain a value of relative expression of each sample relativeto median of the colon samples, as shown in FIG. 45A. The normalizedquantity of each RT sample was then divided by the median of thequantities of the lung samples (sample numbers 26, 29 and 30, Table 2above), to obtain a value of relative expression of each sample relativeto median of the lung samples, as shown in FIG. 45B.

For blood panel—The normalized quantity of each RT sample was thendivided by the median of the quantities of the kidney normal samples(sample numbers 65-67, Table 1 above), to obtain a value of relativeexpression of each sample relative to median of the kidney normalsamples.

The results of this analysis are depicted in the histogram in FIG. 46.Expression of the above-indicated C1ORF32 transcript is high in onelymphoma sample (sample no. 33, Table 1) but in normal brain sample too.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: H19011 junc8-10seg13F1 (SEQ ID NO: 239) forwardprimer; and H19011 junc8-10seg13R1 (SEQ ID NO: 240) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: H19011junc8-10seg13F1R1 (SEQ ID NO: 241).

Forward Primer >H19011_junc8-10seg13F1 (SEQ ID NO: 239)TGTGGAGATTATGCCAGAGTGG Reverse Primer >H19011_junc8-10seg13R1(SEQ ID NO: 240) GACATTTCTCTCGATCGCTCTGTAmplicon >H19011_junc8-10seg13F1R1 (SEQ ID NO: 241)TGTGGAGATTATGCCAGAGTGGGTGTTTGTTGGCCTGGTGCTCCTGGGCGTCTTCCTCTTCTTCGTCCTGGTGGGGATCTGCTGGTGCCAGTGCTGCCCTCACAGCTGCTGCTGCTATGTCCGCTGCCCATGCTGCCCAGATTCCTGCTGGTGCCCTCAAGCCTGTGAGTACAGTGACCGCTGGGGAGACAGAGCG ATCGAGAGAAATGTC

Expression of C1ORF32, chromosome 1 open reading frame 32, H19011transcripts which are detectable by amplicon as depicted in sequencename H19011_junc6-10 (SEQ ID NO: 244) in normal and cancerous lungtissues and in normal and cancerous Colon tissues

Expression of C1ORF32 transcripts detectable by or according tojunc6-10-H19011_junc6-10F1R1 (SEQ ID NO: 244) amplicon and primersH19011_junc6-10F1 (SEQ ID NO: 242) and H19011_junc6-10R1 (SEQ ID NO:243) was measured by real time PCR on lung panel and colon panel. Thesamples used are detailed in Table 3 and Table 5 above, respectively.For each RT sample, the expression of the above amplicon was normalizedto the normalization factor calculated from the expression of severalhouse keeping genes as described in Example 1.

Lung panel—The normalized quantity of each RT sample was then divided bythe median of the quantities of the normal samples (sample numbers51-64, 69 and 70, Table 3 above). Then the reciprocal of this ratio wascalculated, to obtain a value of fold down-regulation for each samplerelative to median of the normal samples.

FIG. 47 is a histogram showing down regulation of the above-indicatedC1ORF32 transcripts in cancerous Lung samples relative to the normalsamples.

As is evident from FIG. 47, the expression of C1ORF32 transcriptsdetectable by the above amplicon in non-small cell carcinoma samples,adenocarcinoma and squamous cell carcinoma was significantly lower thanin the non-cancerous samples (sample numbers 51-64, 69 and 70, Table 3above). Notably down regulation of at least 5 fold was found in 23 outof 39 non-small cell carcinoma samples especially in 8 out of 17adenocarcinoma samples and in 12 out of 16 squamous cell carcinomasamples.

Statistical analysis was applied to verify the significance of theseresults, as described below. The P value for the difference in theexpression levels of C1ORF32 transcripts detectable by the aboveamplicon lung non-small cell carcinoma, lung adenocarcinoma and lungsquamous cell carcinoma samples, versus the normal tissue samples wasdetermined by T test as 1.18e-003, 2.87e-002 and 3.55e-004,respectively.

Threshold of 5 fold down regulation was found to differentiate betweenlung non-small cell carcinoma, lung adenocarcinoma and lung squamouscell carcinoma samples and normal samples with P value of 1.59e-003,3.54e-002 and 4.78e-004, respectively, as checked by exact Fisher test.

The above values demonstrate statistical significance of the results.

Colon panel—The normalized quantity of each RT sample was then dividedby the median of the quantities of the normal samples (sample numbers42-70, Table 5 above). Then the reciprocal of this ratio was calculated,to obtain a value of fold down-regulation for each sample relative tomedian of the normal samples.

FIG. 48 is a histogram showing down regulation of the above-indicatedC1ORF32 transcripts in cancerous Colon samples relative to the normalsamples.

As is evident from FIG. 48, the expression of C1ORF32 transcriptsdetectable by the above amplicon in cancer samples was significantlylower than in the non-cancerous samples (sample numbers 42-70, Table 5above). Notably down regulation of at least 9 fold was found in 23 outof 55 adenocarcinoma samples.

Statistical analysis was applied to verify the significance of theseresults, as described below.

Threshold of 9 fold down regulation was found to differentiate betweencancer and normal samples with P value of 7.39e-006 as checked by exactFisher test.

The above values demonstrate statistical significance of the results.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: H19011 junc6-10F1 (SEQ ID NO: 242) forward primer;and H19011 junc6-10R1 (SEQ ID NO: 243) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: H19011 junc6-10F1R1(SEQ ID NO: 244).

Forward Primer >H19011_junc6-10F1 (SEQ ID NO: 242)ACTCTATTACTGTATTATCACCACCCCAG Reverse Primer >H19011_junc6-10R1(SEQ ID NO: 243) CCAACAAACACCCACTCCAAC Amplicon >H19011_junc6-10F1R1(SEQ ID NO: 244) ACTCTATTACTGTATTATCACCACCCCAGATGACCTGGAGGGGAAAAATGAGGGCTCACTGGGACTGCTGGTGTTGGAGTGGGTGTTTGTTGG

Example 7 Description for Cluster R31375

The present invention relates to a specific antigen FXYD3 and relateddiagnostic and therapeutics based thereon.

According to the present invention, Cluster R31375 (internal ID72360301) features 19 transcripts and 4 segments of interest, the namesfor which are given in Tables 105 and 106, respectively. The selectedprotein variants are given in table 107.

TABLE 105 Transcripts of interest Transcript Name R31375_T0 (SEQ ID NO:51) R31375_T1 (SEQ ID NO: 52) R31375_T2 (SEQ ID NO: 53) R31375_T3 (SEQID NO: 54) R31375_T4 (SEQ ID NO: 55) R31375_T5 (SEQ ID NO: 56) R31375_T6(SEQ ID NO: 57) R31375_T7 (SEQ ID NO: 58) R31375_T8 (SEQ ID NO: 59)R31375_T9 (SEQ ID NO: 60) R31375_T10 (SEQ ID NO: 61) R31375_T11 (SEQ IDNO: 62) R31375_T12 (SEQ ID NO: 63) R31375_T13 (SEQ ID NO: 64) R31375_T19(SEQ ID NO: 65) R31375_T25 (SEQ ID NO: 66) R31375_T26 (SEQ ID NO: 67)R31375_T29 (SEQ ID NO: 68) R31375_T39 (SEQ ID NO: 69)

TABLE 106 Segments of interest Segment Name R31375_N30 (SEQ ID NO: 134)R31375_N33 (SEQ ID NO: 135) R31375_N34 (SEQ ID NO: 136) R31375_N37 (SEQID NO: 137)

TABLE 107 Proteins of interest Protein Name Corresponding TranscriptsR31375_P0 (SEQ R31375_T0 (SEQ ID NO: 51); R31375_T1 (SEQ ID NO: 70) IDNO: 52); R31375_T10 (SEQ ID NO: 61); R31375_T11 (SEQ ID NO: 62);R31375_T12 (SEQ ID NO: 63); R31375_T13 (SEQ ID NO: 64); R31375_T2 (SEQID NO: 53); R31375_T3 (SEQ ID NO: 54); R31375_T4 (SEQ ID NO: 55);R31375_T5 (SEQ ID NO: 56); R31375_T6 (SEQ ID NO: 57); R31375_T7 (SEQ IDNO: 58); R31375_T8 (SEQ ID NO: 59); R31375_T9 (SEQ ID NO: 60) R31375_P14(SEQ R31375_T19 (SEQ ID NO: 65); R31375_T25 ID NO: 72) (SEQ ID NO: 66);R31375_T26 (SEQ ID NO: 67) R31375_P31 (SEQ R31375_T29 (SEQ ID NO: 68) IDNO: 73) R31375_P33 (SEQ R31375_T39 (SEQ ID NO: 69) ID NO: 74)

These sequences are variants of the known protein FXYD domain-containingion transport regulator 3 precursor (SwissProt accession identifierFXYD3_HUMAN (SEQ ID NO: 70); known also according to the synonymsChloride conductance inducer protein Mat-8; Mammary tumor 8 kDa protein;Phospholemman-like), referred to herein as the previously known protein.

FXYD3 was previously identified within a set of genes induced by the neuor Ha-Ras oncogenes in murine breast tumors, and was named Mat-8(Mammary tumor, 8 kDa) (Morrison et al 1995). In normal tissues, FXYD3is mainly expressed in the uterus, stomach, colon and skin (Morrison etal 1995). Its expression was found elevated in human primary breasttumors, as well as prostate carcinoma and pancreatic ductaladenocarcinoma (Grzmil et al 2004, Kayed et al 2006). Specificinhibition of its expression by siRNA in prostate cancer cell linesindicates a role in cellular proliferation (Grzmil et al 2004).

FXYD3 belongs to the FXYD family proteins. The seven known members ofthis family are all small membrane proteins that contain a commonsignature of 6 amino acids comprising the FXYD motif. Most FXYDproteins, including human FXYD3, are type I membrane proteins,containing a transmembrane domain and a cleavable signal peptide.However, the signal peptide of mouse FXYD3 is not cleaved and the signalpeptide may act as a second transmembrane domain (Crambert et al 2005,Geering 2006). FXYD3, like other members of the FXYD family, interactswith Na/K-ATPase and modulates its activity in a tissue-specific manner(Crambert et al 2005, Arimochi et al 2007, Geering 2006).

Two splice variant isoforms of FXYD3 have been previously described(Bibert et al 2006). These differ in a 26 amino acids in frame insertionafter the transmembrane domain, and are differentially expressed duringcell differentiation. Furthermore, both isoforms are able to stablyassociate with Na/K-ATPase and play different functional roles in theregulation of the activities of this ATPase (Bibert et al 2006).

In addition, WO 2003101283 may be relevant to the present invention.This PCT application purports to disclose that R31375_P0 (SEQ ID NO:70)(wild-type FXYD3 nucleic acid coding sequence reported herein) is adifferentially expressed nucleic acid which encodes a protein sequencesthat allegedly may be used as a diagnostic marker for human lung cancer.

Further, WO2003000012 purports to disclose a human breast cancer relatedprotein referred to as protein #12 that seems to correspond to theR31375_ P0 (wild-type) disclosed herein. Also, U.S. Pat. No. 7,189,507discloses a gene referred to as MAT8 in a long table of gene sequencesthat seems to correspond to R31375_P0 (SEQ ID NO:70). The table seems tosuggest that this gene may be expressed in ovarian cancer.

Protein FXYD domain-containing ion transport regulator 3 precursor (SEQID NO:70) is known or believed to have the following functions: Inducesa hyperpolarization-activated chloride current when expressed in Xenopusoocytes. May be a modulator capable of activating endogenous oocytechannels. Known polymorphisms for this sequence are as shown in Table108.

TABLE 108 Amino acid mutations for Known Protein SNP positions onamino acid sequence Comment 36-37 Missing 58 S                        ->SEWRSSGEQAGRGWGSPPLTTQLSPTG

Protein FXYD domain-containing ion transport regulator 3 precursor (SEQID NO:70) localization is believed to be Membrane; single-pass type Imembrane protein (Potential).

The following GO Annotations apply to the previously known protein. Thefollowing annotations were found: chloride transport, which areannotations related to Biological Process; chloride channel activity,which are annotations related to Molecular Function; and integral toplasma membrane, which are annotations related to Cellular Component.

The GO assignment relies on information from one or more of theSwissProt/TremBl Protein knowledge base, available from<http://www.expasy.ch/sprot/>; or Locuslink, available from<http://www.ncbi.nlm.nih.gov/projects/LocusLink/>.

According to the present invention, novel FXYD3 splice variants wereidentified as membrane bound proteins that are predicted to be overexpressed in cancers. FXYD3 is a type I membrane bound protein (Bibertet. alAl 2006) and according to the present invention 3 novel splicevariants of this protein are provided. The novel splice variant referredherein as R31375_P14 (SEQ ID NO:72) has an additional in-frame exon inits extracellular region. The addition of this exon increases the lengthof the extracellular region by 298 amino acids, which comprises asignificant addition to rather short extracellular domain of 18 aminoacids of the wild type protein. The novel splice variant referred hereinas R31375_P31 (SEQ ID NO:73) has an additional in-frame exon in thejuxtamembrane domain of FXYD3, which adds 26 new amino acids to theintracellular region. The novel splice variant referred herein asR31375_P33 (SEQ ID NO:74) skips the 3rd coding exon of the wild typeFXYD3 and, like R31375_P31 (SEQ ID NO:73), has the additional in-frameexon in the juxtamembrane domain. This causes the deletion of 8 aminoacids in the ectodomain.

According to the present invention FXYD3 and R31375_P14 (SEQ ID NO:72)were shown to be overexpressed in ovarian cancer.

As noted above, cluster R31375 features 19 transcripts, which werelisted in Table 105 above. These transcripts encode for proteins whichare variants of protein FXYD domain-containing ion transport regulator 3precursor (SEQ ID NO:70). A description of each variant proteinaccording to the present invention is now provided.

Variant protein R31375_P0 (SEQ ID NO:70) according to the presentinvention has an amino acid sequence encoded by transcripts R31375_T0(SEQ ID NO:51), R31375_T1 (SEQ ID NO:52), R31375_T10 (SEQ ID NO:61),R31375_T11 (SEQ ID NO:62), R31375_T12 (SEQ ID NO:63), R31375_T13 (SEQ IDNO:64), R31375_T2 (SEQ ID NO:53), R31375_T3 (SEQ ID NO:54), R31375_T4(SEQ ID NO:55), R31375_T5 (SEQ ID NO:56), R31375_T6 (SEQ ID NO:57),R31375_T7 (SEQ ID NO:58), R31375_T8 (SEQ ID NO:59) and R31375_T9 (SEQ IDNO:60).

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein R31375_P0 (SEQ ID NO:70) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table109, (given according to their positions on the amino acid sequence,with the alternative amino acids listed (SEQ ID NO:70)).

TABLE 109 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 3 K −> R 19 D −> A 19 D −> V 50 A −> P 75 E −>

The coding portion of transcript R31375_T0 (SEQ ID NO:51) starts atposition 491 and ends at position 751. The transcript also has thefollowing SNPs as listed in Table 110 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 110 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> A 469 A −> G 498, 900, 1571 A −> T 546 A −> C 546 T −> A607, 926  T −> G 607 G −> C 638 G −> 713 C −> T 792 G −> A 901, 1572

The coding portion of transcript R31375_T1 (SEQ ID NO:52) starts atposition 795 and ends at position 1055. The transcript also has thefollowing SNPs as listed in Table 111 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 111 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence G −> A 513, 1205, 1876 C −> A 773 A −> G 802, 1204, 1875 A −> T850 A −> C 850 T −> A 911, 1230 T −> G 911 G −> C 942 G −> 1017  C −> T1096 

The coding portion of transcript R31375_T10 (SEQ ID NO:61) starts atposition 1826 and ends at position 2086. The transcript also has thefollowing SNPs as listed in Table 112 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 112 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> A  469 A −> G 639, 984, 1504, 1833, 2235, 2906 A −> C 860,1183, 1447, 1504, 1881 C −> 1063 A −> T 1183, 1447, 1881 T −> A 1942,2261 T −> G 1942 G −> C 1973 G −> 2048 C −> T 2127 G −> A 2236, 2907

The coding portion of transcript R31375_T11 (SEQ ID NO:62) starts atposition 613 and ends at position 873. The transcript also has thefollowing SNPs as listed in Table 113 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 113 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> A 469 A −> G 620, 1022, 1693 A −> T 668 A −> C 668 T −> A 729, 1048 T −> G 729 G −> C 760 G −> 835 C −> T 914 G −> A 1023, 1694

The coding portion of transcript R31375_T12 (SEQ ID NO:63) starts atposition 711 and ends at position 971. The transcript also has thefollowing SNPs as listed in Table 114 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 114 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> A 469 G −> 545, 933 G −> C 545, 858 C −> 549 A −> G 718,1120, 1791 A −> T 766 A −> C 766 T −> A  827, 1146 T −> G 827 C −> T1012  G −> A 1121, 1792

The coding portion of transcript R31375_T13 (SEQ ID NO:64) starts atposition 1015 and ends at position 1275. The transcript also has thefollowing SNPs as listed in Table 115 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 115 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence G −> A 513, 1425, 2096 C −> A  773 G −> 849, 1237 G −> C 849,1162 C −>  853 A −> G 1022, 1424, 2095 A −> T 1070 A −> C 1070 T −> A1131, 1450  T −> G 1131 C −> T 1316

The coding portion of transcript R31375_T2 (SEQ ID NO:53) starts atposition 678 and ends at position 938. The transcript also has thefollowing SNPs as listed in Table 116 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 116 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence G −> A 513, 1088, 1759 C −> A 656 A −> G 685, 1087, 1758 A −> T733 A −> C 733 T −> A 794, 1113 T −> G 794 G −> C 825 G −> 900 C −> T979

The coding portion of transcript R31375_T3 (SEQ ID NO:54) starts atposition 572 and ends at position 832. The transcript also has thefollowing SNPs as listed in Table 117 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 117 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> A 550 A −> G 579, 981, 1652 A −> T 627 A −> C 627 T −> A688, 1007 T −> G 688 G −> C 719 G −> 794 C −> T 873 G −> A 982, 1653

The coding portion of transcript R31375_T4 (SEQ ID NO:55) starts atposition 575 and ends at position 835. The transcript also has thefollowing SNPs as listed in Table 118 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 118 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> A 469 A −> G 582, 984, 1655 A −> T 630 A −> C 630 T −> A691, 1010 T −> G 691 G −> C 722 G −> 797 C −> T 876 G −> A 985, 1656

The coding portion of transcript R31375_T5 (SEQ ID NO:56) starts atposition 656 and ends at position 916. The transcript also has thefollowing SNPs as listed in Table 119 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 119 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> A 550 A −> G 663, 1065, 1736 A −> T 711 A −> C 711 T −> A 772, 1091 T −> G 772 G −> C 803 G −> 878 C −> T 957 G −> A 1066, 1737

The coding portion of transcript R31375_T6 (SEQ ID NO:57) starts atposition 697 and ends at position 957. The transcript also has thefollowing SNPs as listed in Table 120 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 120 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> A 469 A −> G 704, 1106, 1777 A −> T 752 A −> C 752 T −> A 813, 1132 T −> G 813 G −> C 844 G −> 919 C −> T 998 G −> A 1107, 1778

The coding portion of transcript R31375_T7 (SEQ ID NO:58) starts atposition 2475 and ends at position 2735. The transcript also has thefollowing SNPs as listed in Table 121 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 121 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> A  469 A −> G 639, 984, 1504, 2482, 2884, 3555 A −> C 860,1183, 1447, 1504, 2530 C −> 1063 A −> T 1183, 1447, 2530 C −> T 1999,2022, 2776 −> G 2279 −> A 2280 G −> T 2285 G −> C 2285, 2622 T −> A2591, 2910 T −> G 2591 G −> 2697 G −> A 2885, 3556

The coding portion of transcript R31375_T8 (SEQ ID NO:59) starts atposition 1329 and ends at position 1589. The transcript also has thefollowing SNPs as listed in Table 122 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 122 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> A  469 A −> G 639, 984, 1336, 1738, 2409 A −> C 860, 1183,1384 C −> 1063 A −> T 1183, 1384 T −> A 1445, 1764 T −> G 1445 G −> C1476 G −> 1551 C −> T 1630 G −> A 1739, 2410

The coding portion of transcript R31375_T9 (SEQ ID NO:60) starts atposition 2586 and ends at position 2846. The transcript also has thefollowing SNPs as listed in Table 123 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 123 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> A 469, 2531, 2549 A −> G 639, 984, 1504, 2593, 2995, 3666A −> C 860, 1183, 1447, 1504, 2641 C −> 1063 A −> T 1183, 1447, 2641 C−> T 1999, 2022, 2887 −> G 2279 −> A 2280 G −> T 2285 G −> C 2285, 2733C −> G 2531, 2549 G −> A 2550, 2996, 3667 T −> A 2702, 3021 T −> G 2702G −> 2808

Variant protein R31375_P14 (SEQ ID NO:72) according to the presentinvention has an amino acid sequence as encoded by transcriptsR31375_T19 (SEQ ID NO:65), R31375_T25 (SEQ ID NO:66) and R31375_T26 (SEQID NO:67). Alignments to one or more previously published proteinsequences are shown in FIG. 49A. A brief description of the relationshipof the variant protein according to the present invention to each suchaligned protein is as follows:

1. Comparison report between R31375_P14 (SEQ ID NO:72) and knownproteins FXYD3_HUMAN, NP_(—)005962 and Q6IB59_HUMAN (SEQ ID NO: 70)(FIG. 49A):

A. An isolated chimeric polypeptide encoding for R31375_P14 (SEQ IDNO:72), comprising a first amino acid sequence being at least 90%homologous to MQKVTLGLLVFLAGFPVLDANDLEDKNSPFYY corresponding to aminoacids 1-32 of known proteins FXYD3_HUMAN, NP_(—)005962 and Q6IB59_HUMAN(SEQ ID NO: 70), which also corresponds to amino acids 1-32 ofR31375_P14 (SEQ ID NO:72), a second amino acid sequence being at least70%, optionally at least 80%, preferably at least 85%, more preferablyat least 90% and most preferably at least 95, 96, 97, 98 or 99%homologous to a polypeptide having the sequenceGAPYIFVKRMGGQMKRTQAGTEVPSTFLL (SEQ ID NO: 294) corresponding to aminoacids 33-61 of R31375_P14 (SEQ ID NO:72), and a third amino acidsequence being at least 90% homologous toDWHSLQVGGLICAGVLCAMGIIIVMSAKCKCKFGQKSGHHPGETPPLITPGSAQ S correspondingto amino acids 33-87 of known proteins FXYD3_HUMAN, NP_(—)005962 andQ6IB59_HUMAN (SEQ ID NO: 70), which also corresponds to amino acids62-116 of R31375_P14 (SEQ ID NO:72), wherein said first amino acidsequence, second amino acid sequence and third amino acid sequence arecontiguous and in a sequential order.

B. An isolated polypeptide encoding for an edge portion of R31375_P14(SEQ ID NO:72), comprising an amino acid sequence being at least 70%,optionally at least about 80%, preferably at least about 85%, morepreferably at least about 90% and most preferably at least about 95, 96,97, 98 or 99% homologous to the sequence GAPYIFVKRMGGQMKRTQAGTEVPSTFLL(SEQ ID NO: 294) of R31375_P14 (SEQ ID NO:72).

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein R31375_P14 (SEQ ID NO:72) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table124, (given according to their positions on the amino acid sequence,with the alternative amino acids listed (SEQ ID NO:72)).

TABLE 124 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 3 K −> R 19 D −> A 19 D −> V 46 M −> I 47 K −> E 79A −> P 104 E −>

The coding portion of transcript R31375_T19 (SEQ ID NO:65) starts atposition 491 and ends at position 838. The transcript also has thefollowing SNPs as listed in Table 125 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 125 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> A 469 A −> G 498, 629, 987, 1658 A −> T 546 A −> C 546 G−> A 628, 988, 1659 T −> A 694, 1013 T −> G 694 G −> C 725 G −> 800 C −>T 879

The coding portion of transcript R31375_T25 (SEQ ID NO:66) starts atposition 575 and ends at position 922. The transcript also has thefollowing SNPs as listed in Table 126 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 126 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> A 469 A −> G 582, 713, 1071, 1742 A −> T 630 A −> C 630 G−> A 712, 1072, 1743 T −> A 778, 1097 T −> G 778 G −> C 809 G −> 884 C−> T 963

The coding portion of transcript R31375_T26 (SEQ ID NO:67) starts atposition 1443 and ends at position 1790. The transcript also has thefollowing SNPs as listed in Table 127 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 127 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> A  469 A −> G 639, 984, 1450, 1581, 1939, 2610 A −> C 860,1183, 1498 C −> 1063 A −> T 1183, 1498 G −> A 1580, 1940, 2611 T −> A1646, 1965 T −> G 1646 G −> C 1677 G −> 1752 C −> T 1831

Variant protein R31375_P31 (SEQ ID NO:73) according to the presentinvention has an amino acid sequence as encoded by transcript R31375_T29(SEQ ID NO:68). Alignments to one or more previously published proteinsequences are given in FIGS. 49B and 49C. A brief description of therelationship of the variant protein according to the present inventionto each such aligned protein is as follows:

1. Comparison report between R31375_P31 (SEQ ID NO:73) and knownproteins FXYD3_HUMAN, NP_(—)005962 and Q6IB59_HUMAN (SEQ ID NO: 70)(FIG. 49B):

A. An isolated chimeric polypeptide encoding for R31375_P31 (SEQ IDNO:73), comprising a first amino acid sequence being at least 90%homologous to MQKVTLGLLVFLAGFPVLDANDLEDKNSPFYYDWHSLQVGGLICAGVLCAMGIIIVMS corresponding to amino acids 1-58 of known proteins FXYD3_HUMAN,NP_(—)005962 and Q6IB59_HUMAN (SEQ ID NO: 70), which also corresponds toamino acids 1-58 of R31375_P31 (SEQ ID NO:73), a second amino acidsequence being at least 70%, optionally at least 80%, preferably atleast 85%, more preferably at least 90% and most preferably at least 95,96, 97, 98 or 99% homologous to a polypeptide having the sequenceEWRSSGEQAGRGWGSPPLTTQLSPTG (SEQ ID NO: 295) corresponding to amino acids59-84 of R31375_P31 (SEQ ID NO:73), and a third amino acid sequencebeing at least 90% homologous to AKCKCKFGQKSG corresponding to aminoacids 59-70 of known proteins FXYD3_HUMAN, NP_(—)005962 and Q6IB59_HUMAN(SEQ ID NO: 70), which also corresponds to amino acids 85-96 ofR31375_P31 (SEQ ID NO:73), wherein said first amino acid sequence,second amino acid sequence and third amino acid sequence are contiguousand in a sequential order.

B. An isolated polypeptide encoding for an edge portion of R31375_P31(SEQ ID NO:73), comprising an amino acid sequence being at least 70%,optionally at least about 80%, preferably at least about 85%, morepreferably at least about 90% and most preferably at least about 95, 96,97, 98 or 99% homologous to the sequence EWRSSGEQAGRGWGSPPLTTQLSPTG (SEQID NO: 295) of R31375_P31 (SEQ ID NO:73).

2. Comparison report between R31375_P31 (SEQ ID NO:73) and knownproteins NP_(—)068710 (SEQ ID NO: 71) (FIG. 49C):

A. An isolated chimeric polypeptide encoding for R31375_P31 (SEQ IDNO:73), comprising a amino acid sequence being at least 90% homologousto MQKVTLGLLVFLAGFPVLDANDLEDKNSPFYYDWHSLQVGGLICAGVLCAMGIIIVMSEWRSSGEQAGRGWGSPPLTTQLSPTGAKCKCKFGQKSG corresponding to amino acids1-96 of known proteins NP_(—)068710 (SEQ ID NO: 71), which alsocorresponds to amino acids 1-96 of R31375_P31 (SEQ ID NO:73), whereinsaid and first amino acid sequence are contiguous and in a sequentialorder.

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: membrane.

Variant protein R31375_P31 (SEQ ID NO:73) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table128, (given according to their positions on the amino acid sequence,with the alternative amino acids listed (SEQ ID NO:73)).

TABLE 128 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 3 K −> R 19 D −> A 19 D −> V 50 A −> P

Variant protein R31375_P31 (SEQ ID NO:73) is encoded by the transcriptR31375_T29 (SEQ ID NO:68), for which the coding portion starts atposition 491 and ends at position 778. The transcript also has thefollowing SNPs as listed in Table 129 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 129 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> A 469 A −> G 498, 1294, 1965 A −> T 546 A −> C 546 T −> A 607, 1320 T −> G 607 G −> C 638 G −> 1107  C −> T 1186  G −> A 1295,1966

Variant protein R31375_P33 (SEQ ID NO:74) according to the presentinvention has an amino acid sequence as encoded by transcript R31375_T39(SEQ ID NO:69). Alignments to one or more previously published proteinsequences are given in FIGS. 49D and 49E. A brief description of therelationship of the variant protein according to the present inventionto each such aligned protein is as follows:

1. Comparison report between R31375_P33 (SEQ ID NO:74) and knownproteins FXYD3_HUMAN, NP_(—)005962 and Q6IB59_HUMAN (SEQ ID NO: 70)(FIG. 49D):

A. An isolated chimeric polypeptide encoding for R31375_P33 (SEQ IDNO:74), comprising a first amino acid sequence being at least 90%homologous to MQKVTLGLLVFLAGFPVLDANDLE corresponding to amino acids 1-24of known proteins FXYD3_HUMAN, NP_(—)005962 and Q6IB59_HUMAN (SEQ ID NO:70), which also corresponds to amino acids 1-24 of R31375_P33 (SEQ IDNO:74), a second amino acid sequence being at least 90% homologous toDWHSLQVGGLICAGVLCAMGIIIVMS corresponding to amino acids 33-58 of knownproteins FXYD3_HUMAN, NP_(—)005962 and Q6IB59_HUMAN (SEQ ID NO: 70),which also corresponds to amino acids 25-50 of R31375_P33 (SEQ IDNO:74), a third amino acid sequence being at least 70%, optionally atleast 80%, preferably at least 85%, more preferably at least 90% andmost preferably at least 95, 96, 97, 98 or 99% homologous to apolypeptide having the sequence EWRSSGEQAGRGWGSPPLTTQLSPTG (SEQ ID NO:295) corresponding to amino acids 51-76 of R31375_P33 (SEQ ID NO:74),and a fourth amino acid sequence being at least 90% homologous toAKCKCKFGQKSG corresponding to amino acids 59-70 of known proteinsFXYD3_HUMAN, NP_(—)005962 and Q6IB59_HUMAN (SEQ ID NO: 70), which alsocorresponds to amino acids 77-88 of R31375_P33 (SEQ ID NO:74), whereinsaid first amino acid sequence, second amino acid sequence, third aminoacid sequence and fourth amino acid sequence are contiguous and in asequential order.

B. An isolated chimeric polypeptide encoding for an edge portion ofR31375_P33 (SEQ ID NO:74), comprising a polypeptide having a length “n”,wherein n is at least about 10 amino acids in length, optionally atleast about 20 amino acids in length, preferably at least about 30 aminoacids in length, more preferably at least about 40 amino acids in lengthand most preferably at least about 50 amino acids in length, wherein atleast two amino acids comprise ED, having a structure as follows: asequence starting from any of amino acid numbers 24−x to 24; and endingat any of amino acid numbers 25+((n−2)−x), in which x varies from 0 ton−2.

C. An isolated polypeptide encoding for an edge portion of R31375_P33(SEQ ID NO:74), comprising an amino acid sequence being at least 70%,optionally at least about 80%, preferably at least about 85%, morepreferably at least about 90% and most preferably at least about 95, 96,97, 98 or 99% homologous to the sequence EWRSSGEQAGRGWGSPPLTTQLSPTG (SEQID NO: 295) of R31375_P33 (SEQ ID NO:74).

2. Comparison report between R31375_P33 (SEQ ID NO:74) and knownproteins NP_(—)068710 (SEQ ID NO: 71) (FIG. 49E):

A. An isolated chimeric polypeptide encoding for R31375_P33 (SEQ IDNO:74), comprising a first amino acid sequence being at least 90%homologous to MQKVTLGLLVFLAGFPVLDANDLE corresponding to amino acids 1-24of known proteins NP_(—)068710 (SEQ ID NO: 71), which also correspondsto amino acids 1-24 of R31375_P33 (SEQ ID NO:74), and a second aminoacid sequence being at least 90% homologous toDWHSLQVGGLICAGVLCAMGIIIVMSEWRSSGEQAGRGWGSPPLTTQLSPTGAK CKCKFGQKSGcorresponding to amino acids 33-96 of known proteins NP_(—)068710 (SEQID NO: 71), which also corresponds to amino acids 25-88 of R31375_P33(SEQ ID NO:74), wherein said first amino acid sequence and second aminoacid sequence are contiguous and in a sequential order.

B. An isolated chimeric polypeptide encoding for an edge portion ofR31375_P33 (SEQ ID NO:74), comprising a polypeptide having a length “n”,wherein n is at least about 10 amino acids in length, optionally atleast about 20 amino acids in length, preferably at least about 30 aminoacids in length, more preferably at least about 40 amino acids in lengthand most preferably at least about 50 amino acids in length, wherein atleast two amino acids comprise ED, having a structure as follows: asequence starting from any of amino acid numbers 24−x to 24; and endingat any of amino acid numbers 25+((n−2)−x), in which x varies from 0 ton−2.

The localization of the variant protein was determined according toresults from a number of different software programs and analyses,including analyses from SignalP and other specialized programs. Thevariant protein is believed to be located as follows with regard to thecell: secreted.

Variant protein R31375_P33 (SEQ ID NO:74) also has the followingnon-silent SNPs (Single Nucleotide Polymorphisms) as listed in Table130, (given according to their positions on the amino acid sequence,with the alternative amino acids listed (SEQ ID NO:74)).

TABLE 130 Amino acid mutations SNP positions on Alternative amino aminoacid sequence acids 3 K −> R 19 D −> A 19 D −> V 42 A −> P

The coding portion of transcript R31375_T39 (SEQ ID NO:69) starts atposition 491 and ends at position 754. The transcript also has thefollowing SNPs as listed in Table 131 (given according to their positionon the nucleotide sequence, with the alternative nucleic acid listed).

TABLE 131 Nucleic acid SNPs SNP positions on Polymorphism nucleotidesequence C −> A 469 A −> G 498, 1270, 1941 A −> T 546 A −> C 546 T −> A 583, 1296 T −> G 583 G −> C 614 G −> 1083  C −> T 1162  G −> A 1271,1942

According to an optional embodiment of the present invention, shortsegments related to the above cluster are also provided. These segmentsare up to about 120 bp in length, and so are included in a separatedescription.

Segment cluster R31375_N30 (SEQ ID NO:134) according to the presentinvention is supported by 7 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: R31375_T19 (SEQ ID NO:65), R31375_T25 (SEQ IDNO:66) and R31375_T26 (SEQ ID NO:67). Table 132 below describes thestarting and ending position of this segment on each transcript.

TABLE 132 Segment location on transcripts Segment Segment Transcriptname starting position ending position R31375_T19 (SEQ ID NO: 65) 588674 R31375_T25 (SEQ ID NO: 66) 672 758 R31375_T26 (SEQ ID NO: 67) 15401626

Segment cluster R31375_N33 (SEQ ID NO:135) according to the presentinvention is supported by 278 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: R31375_TO (SEQ ID NO:51), R31375_T1 (SEQ IDNO:52), R31375_T10 (SEQ ID NO:61), R31375_T11 (SEQ ID NO:62), R31375_T12(SEQ ID NO:63), R31375_T13 (SEQ ID NO:64), R31375_T19 (SEQ ID NO:65),R31375_T2 (SEQ ID NO:53), R31375_T25 (SEQ ID NO:66), R31375_T26 (SEQ IDNO:67), R31375_T29 (SEQ ID NO:68), R31375_T3 (SEQ ID NO:54), R31375_T39(SEQ ID NO:69), R31375_T4 (SEQ ID NO:55), R31375_T5 (SEQ ID NO:56),R31375_T6 (SEQ ID NO:57), R31375_T7 (SEQ ID NO:58), R31375_T8 (SEQ IDNO:59) and R31375_T9 (SEQ ID NO:60). Table 133 below describes thestarting and ending position of this segment on each transcript.

TABLE 133 Segment location on transcripts Segment Segment Transcriptname starting position ending position R31375_T0 (SEQ ID NO: 51) 588 631R31375_T1 (SEQ ID NO: 52) 892 935 R31375_T10 (SEQ ID NO: 61) 1923 1966R31375_T11 (SEQ ID NO: 62) 710 753 R31375_T12 (SEQ ID NO: 63) 808 851R31375_T13 (SEQ ID NO: 64) 1112 1155 R31375_T19 (SEQ ID NO: 65) 675 718R31375_T2 (SEQ ID NO: 53) 775 818 R31375_T25 (SEQ ID NO: 66) 759 802R31375_T26 (SEQ ID NO: 67) 1627 1670 R31375_T29 (SEQ ID NO: 68) 588 631R31375_T3 (SEQ ID NO: 54) 669 712 R31375_T39 (SEQ ID NO: 69) 564 607R31375_T4 (SEQ ID NO: 55) 672 715 R31375_T5 (SEQ ID NO: 56) 753 796R31375_T6 (SEQ ID NO: 57) 794 837 R31375_T7 (SEQ ID NO: 58) 2572 2615R31375_T8 (SEQ ID NO: 59) 1426 1469 R31375_T9 (SEQ ID NO: 60) 2683 2726

Segment cluster R31375_N34 (SEQ ID NO:136) according to the presentinvention is supported by 275 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: R31375_TO (SEQ ID NO:51), R31375_T1 (SEQ IDNO:52), R31375_T10 (SEQ ID NO:61), R31375_T11 (SEQ ID NO:62), R31375_T12(SEQ ID NO:63), R31375_T13 (SEQ ID NO:64), R31375_T19 (SEQ ID NO:65),R31375_T2 (SEQ ID NO:53), R31375_T25 (SEQ ID NO:66), R31375_T26 (SEQ IDNO:67), R31375_T29 (SEQ ID NO:68), R31375_T3 (SEQ ID NO:54), R31375_T39(SEQ ID NO:69), R31375_T4 (SEQ ID NO:55), R31375_T5 (SEQ ID NO:56),R31375_T6 (SEQ ID NO:57), R31375_T7 (SEQ ID NO:58), R31375_T8 (SEQ IDNO:59) and R31375_T9 (SEQ ID NO:60). Table 134 below describes thestarting and ending position of this segment on each transcript.

TABLE 134 Segment location on transcripts Segment Segment Transcriptname starting position ending position R31375_T0 (SEQ ID NO: 51) 632 662R31375_T1 (SEQ ID NO: 52) 936 966 R31375_T10 (SEQ ID NO: 61) 1967 1997R31375_T11 (SEQ ID NO: 62) 754 784 R31375_T12 (SEQ ID NO: 63) 852 882R31375_T13 (SEQ ID NO: 64) 1156 1186 R31375_T19 (SEQ ID NO: 65) 719 749R31375_T2 (SEQ ID NO: 53) 819 849 R31375_T25 (SEQ ID NO: 66) 803 833R31375_T26 (SEQ ID NO: 67) 1671 1701 R31375_T29 (SEQ ID NO: 68) 632 662R31375_T3 (SEQ ID NO: 54) 713 743 R31375_T39 (SEQ ID NO: 69) 608 638R31375_T4 (SEQ ID NO: 55) 716 746 R31375_T5 (SEQ ID NO: 56) 797 827R31375_T6 (SEQ ID NO: 57) 838 868 R31375_T7 (SEQ ID NO: 58) 2616 2646R31375_T8 (SEQ ID NO: 59) 1470 1500 R31375_T9 (SEQ ID NO: 60) 2727 2757

Segment cluster R31375_N37 (SEQ ID NO:137) according to the presentinvention is supported by 254 libraries. The number of libraries wasdetermined as previously described. This segment can be found in thefollowing transcripts: R31375_TO (SEQ ID NO:51), R31375_T1 (SEQ IDNO:52), R31375_T10 (SEQ ID NO:61), R31375_T11 (SEQ ID NO:62), R31375_T12(SEQ ID NO:63), R31375_T13 (SEQ ID NO:64), R31375_T19 (SEQ ID NO:65),R31375_T2 (SEQ ID NO:53), R31375_T25 (SEQ ID NO:66), R31375_T26 (SEQ IDNO:67), R31375_T29 (SEQ ID NO:68), R31375_T3 (SEQ ID NO:54), R31375_T39(SEQ ID NO:69), R31375_T4 (SEQ ID NO:55), R31375_T5 (SEQ ID NO:56),R31375_T6 (SEQ ID NO:57), R31375_T7 (SEQ ID NO:58), R31375_T8 (SEQ IDNO:59) and R31375_T9 (SEQ ID NO:60). Table 135 below describes thestarting and ending position of this segment on each transcript.

TABLE 135 Segment location on transcripts Segment Segment Transcriptname starting position ending position R31375_T0 (SEQ ID NO: 51) 663 699R31375_T1 (SEQ ID NO: 52) 967 1003 R31375_T10 (SEQ ID NO: 61) 1998 2034R31375_T11 (SEQ ID NO: 62) 785 821 R31375_T12 (SEQ ID NO: 63) 883 919R31375_T13 (SEQ ID NO: 64) 1187 1223 R31375_T19 (SEQ ID NO: 65) 750 786R31375_T2 (SEQ ID NO: 53) 850 886 R31375_T25 (SEQ ID NO: 66) 834 870R31375_T26 (SEQ ID NO: 67) 1702 1738 R31375_T29 (SEQ ID NO: 68) 741 777R31375_T3 (SEQ ID NO: 54) 744 780 R31375_T39 (SEQ ID NO: 69) 717 753R31375_T4 (SEQ ID NO: 55) 747 783 R31375_T5 (SEQ ID NO: 56) 828 864R31375_T6 (SEQ ID NO: 57) 869 905 R31375_T7 (SEQ ID NO: 58) 2647 2683R31375_T8 (SEQ ID NO: 59) 1501 1537 R31375_T9 (SEQ ID NO: 60) 2758 2794

Expression of FXYD3 domain containing ion transport regulator 3 R31375transcripts which are detectable by amplicon as depicted in sequencename R31375_junc30-33 (SEQ ID NO: 247) in normal and cancerous Ovarytissues and in different normal tissues

Expression of FXYD3 domain containing ion transport regulator 3transcripts detectable by or according to junc30-33—R31375_junc30-33(SEQ ID NO: 247) amplicon and primers R31375_junc30-33F1 (SEQ ID NO:245) and R31375_junc30-33R1 (SEQ ID NO: 246) was measured by real timePCR on ovary panel and normal panel. The samples used are detailed inTable 4 and Table 2 above, respectively. For each RT sample, theexpression of the above amplicon was normalized to the normalizationfactor calculated from the expression of several house keeping genes asdescribed in Example 1.

Ovary panel—Non-detected samples (samples no. 33 and 53, Table 4), wereassigned Ct value of 41 and were calculated accordingly. The normalizedquantity of each RT sample was then divided by the median of thequantities of the normal samples (sample numbers 52-78, Table 4 above),to obtain a value of fold up-regulation for each sample relative tomedian of the normal samples.

FIG. 50 is a histogram showing over expression of the above-indicatedFXYD3 domain containing ion transport regulator 3 transcripts incancerous Ovary samples relative to the normal samples.

As is evident from FIG. 50, the expression of FXYD3 domain containingion transport regulator 3 transcripts detectable by the above ampliconin adenocarcinoma samples specifically mucinous carcinoma andendometroid samples was significantly higher than in the non-canceroussamples (sample numbers 52-78, Table 4 above). Notably anover-expression of at least 18 fold was found in 13 out of 37adenocarcinoma samples, specifically 7 out of 9 mucinous carcinomasamples and in 4 out of 10 endometroid samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. The P value for the difference in theexpression levels of FXYD3 domain containing ion transport regulator 3transcripts detectable by the above amplicon in Ovary adenocarcinomasamples, mucinous carcinoma samples and endometroid samples versus thenormal tissue samples was determined by T test as 9.75e-004, 1.92e-002and 1.55e-002, respectively.

Threshold of 18 fold over expression was found to differentiate betweenadenocarcinoma mucinous carcinoma samples and endometroid samples andnormal samples with P value of 2.71e-004, 4.31e-006 and 3.18e-003,respectively, as checked by exact Fisher test.

The above values demonstrate statistical significance of the results.

Normal panel—Non-detected samples (samples no. 50 and 54, Table 2) wereassigned Ct value of 41 and were calculated accordingly. The normalizedquantity of each RT sample was then divided by the median of thequantities of the ovary samples (sample numbers 31-34, Table 2 above),to obtain a value of relative expression of each sample relative tomedian of the ovary samples, as shown in FIG. 51.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: R31375_junc30-33F1 (SEQ ID NO: 245) forwardprimer; and R31375_junc30-33R1 (SEQ ID NO: 246) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: R31375_junc30-33 (SEQID NO: 247).

Forward Primer >R31375_junc30-33F1 (SEQ ID NO: 245):GTGCTCCATATATATTTGTCAAGAGAATG Reverse Primer >R31375_junc30-33R1(SEQ ID NO: 246): GGAGGCTGTGCCAGTCTAGG Amplicon >R31375_junc30-33(SEQ ID NO: 247): GTGCTCCATATATATTTGTCAAGAGAATGGGGGGACAGATGAAGAGGACACAGGCTGGCACTGAGGTCCCCTCCACTTTCCTCCTAGACTGGCACAGC CTCC

Expression of FXYD3 domain containing ion transport regulator 3 R31375transcripts which are detectable by amplicon as depicted in sequencename R31375_seg33junc34-37 (SEQ ID NO: 250) in normal and cancerousOvary tissues and in different normal tissues

Expression of FXYD3 domain containing ion transport regulator 3transcripts detectable by or according toseg33junc34-37—R31375_seg33junc34-37 (SEQ ID NO: 250) amplicon andprimers R31375_seg33junc34-37F1 (SEQ ID NO: 248) andR31375_seg33junc34-37R1 (SEQ ID NO: 249) was measured by real time PCRon ovary panel and normal panel. The samples used are detailed in Table4 and Table 2 above, respectively. For each RT sample, the expression ofthe above amplicon was normalized to the normalization factor calculatedfrom the expression of several house keeping genes as described inExample 1.

Ovary panel—Non-detected samples (samples no. 52, 61 and 70, Table 4)were assigned Ct value of 41 and were calculated accordingly. Thenormalized quantity of each RT sample was then divided by the median ofthe quantities of the normal samples (sample numbers 52-78, Table 4above), to obtain a value of fold up-regulation for each sample relativeto median of the normal samples.

FIG. 52 is a histogram showing over expression of the above-indicatedFXYD3 domain containing ion transport regulator 3 transcripts incancerous Ovary samples relative to the normal samples.

As is evident from FIG. 52, the expression of FXYD3 domain containingion transport regulator 3 transcripts detectable by the above ampliconin adenocarcinoma samples—serous carcinoma, mucinous carcinoma andendometroid was significantly higher than in the non-cancerous samples(sample numbers 52-78, Table 4 above). Notably an over-expression of atleast 85 fold was found in 20 out of 37 adenocarcinoma samples,specifically in 7 out of 18 serous carcinoma samples, in 8 out of 9mucinous carcinoma samples and in 5 out of 10 endometroid samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. The P value for the difference in theexpression levels of FXYD3 domain containing ion transport regulator 3transcripts detectable by the above amplicon in Ovary adenocarcinomasamples, serous carcinoma samples, mucinous carcinoma samples andendometriod samples versus the normal tissue samples was determined by Ttest as 1.61e-004, 5.40e-003, 1.49e-002 and 9.08e-003, respectively.

Threshold of 85 fold over expression was found to differentiate betweenadenocarcinoma, serous carcinoma, mucinous carcinoma and endometriod andnormal samples with P value of 8.11e-007, 7.01e-004, 2.97e-007 and5.78e-004, respectively, as checked by exact Fisher test.

The above values demonstrate statistical significance of the results.

Normal panel—The normalized quantity of each RT sample was then dividedby the median of the quantities of the ovary samples (sample numbers31-34, Table 2 above), to obtain a value of relative expression of eachsample relative to median of the ovary samples, as shown in FIG. 53.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: R31375_seg33junc34-37F1 (SEQ ID NO: 248) forwardprimer; and R31375_seg33junc34-37R1 (SEQ ID NO: 249) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: R31375_seg33junc34-37(SEQ ID NO: 250).

Forward Primer >R31375_seg33junc34-37F1 (SEQ ID NO: 248)ACTGGCACAGCCTCCAGG Reverse Primer >R31375_seg33junc34-37R1(SEQ ID NO: 249) CATTTGCATTTTGCACTCATG Amplicon >R31375_seg33junc34-37(SEQ ID NO: 250) ACTGGCACAGCCTCCAGGTTGGCGGGCTCATCTGCGCTGGGGTTCTGTGCGCCATGGGCATCATCATCGTCATGAGTGCAAAATGCAAATG

Expression of FXYD3 domain containing ion transport regulator 3 R31375transcripts which are detectable by amplicon as depicted in sequencename R31375_junc20-22seg30F6R6 (SEQ ID NO: 253) in normal and cancerousovary tissues and in different normal tissues.

Expression of FXYD3 domain containing ion transport regulator 3transcripts detectable by or according tojunc20-22seg30—R31375_junc20-22seg30F6R6 (SEQ ID NO: 253) amplicon andprimers R31375_junc20-22seg30F6 (SEQ ID NO: 251) andR31375_junc20-22seg30R6 (SEQ ID NO: 252) was measured by real time PCRon ovary panel and normal panel. The samples used are detailed in Table4 and Table 2 above, respectively. For each RT sample, the expression ofthe above amplicon was normalized to the normalization factor calculatedfrom the expression of several house keeping genes as described inExample 1.

For ovary panel—Non-detected samples (samples no. 2, 6, 9, 12, 15, 19,21, 24, 32, 34, 38, 45, 53, 56-59, 62, 63, 65-67 and 72-78, Table 4)were assigned Ct value of 41 and were calculated accordingly. Thenormalized quantity of each RT sample was then divided by the median ofthe quantities of the normal samples (sample numbers 52, 53, 56-59, 62,63, 65-67 and 72-78, Table 4 above), to obtain a value of foldup-regulation for each sample relative to median of the normal samples.

FIG. 54 is a histogram showing over expression of the above-indicatedFXYD3 transcripts in cancerous Ovary samples relative to the normalsamples.

As is evident from FIG. 54, the expression of FXYD3 transcriptsdetectable by the above amplicon in adenocarcinoma samples, serouscarcinoma samples, mucinous carcinoma samples and endometroid sampleswas significantly higher than in the non-cancerous samples (samplenumbers 52, 53, 56-59, 62, 63, 65-67 and 72-78, Table 4 above). Notablyan over-expression of at least 14 fold was found in 21 out of 33adenocarcinoma samples, 10 out of 16 serous carcinoma samples, in 5 outof 8 mucinous carcinoma samples and in 6 out of 9 endometroid samples.

Statistical analysis was applied to verify the significance of theseresults, as described below. The P value for the difference in theexpression levels of FXYD3 transcripts detectable by the above ampliconin ovary adenocarcinoma samples versus the normal tissue samples wasdetermined by T test as 3.78e-003.

Threshold of 14 fold over expression was found to differentiate betweenadenocarcinoma, serous carcinoma, mucinous carcinoma and endometriod andnormal samples with P value of 4.21e-005, 5.17e-004, 4.46e-003 and1.73e-003, respectively, as checked by exact Fisher test.

The above values demonstrate statistical significance of the results.

For normal panel—Non-detected samples (samples no. 1, 10, 11, 14, 17,25, 29, 31-34, 38, 39, 46, 47, 49-54, 57, 58, 61-65, 68, 69 and 73,Table 2) were assigned Ct value of 41 and were calculated accordingly.The normalized quantity of each RT sample was then divided by the medianof the quantities of the ovary samples (sample numbers 31-34, Table 2above), to obtain a value of relative expression of each sample relativeto median of the ovary samples, as shown in FIG. 55.

Primer pairs are also optionally and preferably encompassed within thepresent invention; for example, for the above experiment, the followingprimer pair was used as a non-limiting illustrative example only of asuitable primer pair: R31375 junc20-22seg30F6 (SEQ ID NO: 251) forwardprimer; and R31375 junc20-22seg30R6 (SEQ ID NO: 252) reverse primer.

The present invention also preferably encompasses any amplicon obtainedthrough the use of any suitable primer pair; for example, for the aboveexperiment, the following amplicon was obtained as a non-limitingillustrative example only of a suitable amplicon: R31375junc20-22seg30F6R6 (SEQ ID NO: 253).

Forward Primer >R31375_junc20-22seg30F6 (SEQ ID NO: 251)TTGTGTTCCTGGCAGGCTTT Reverse Primer >R31375_junc20-22seg30R6(SEQ ID NO: 252) TCATCTGTCCCCCCATTCTCAmplicon >R31375_junc20-22seg30F6R6 (SEQ ID NO: 253)TTGTGTTCCTGGCAGGCTTTCCTGTCCTGGACGCCAATGACCTAGAAGATAAAAACAGTCCTTTCTACTATGGTGCTCCATATATATTTGTCAAGAGA ATGGGGGGACAGATGA

Example 8 Cloning of Full Length Transcripts Encoding VSIG1, ILDR1,LOC253012, AI216611, C1ORF32, FXYD3 Fused to EGFP

Cloning of Full Length transcripts encoding VSIG1, ILDR1, LOC253012,AI216611, C1ORF32, FXYD3 fused to EGFP was done as described below.

First, EGFP expression vector was constructed and then the VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 open reading frames (ORFs) werecloned. EGFP was subcloned into pIRESpuro3 (Clontech catalog number:631619) as follows: EGFP-N1 vector (Clontech catalog number: 6085-1) wasdigested with NheI and NotI to excise the EGFP gene. The EGFP insert wasthen ligated into pIRESpuro3 (Clontech catalog number: 631619), whichwas previously digested with the same enzymes, in order to obtain theEGFP-pIRESpuro3 vector.

Cloning of the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32, FXYD3 openreading frames (ORFs) was done using the following steps:

1. A reverse transcription reaction was carried out as follows: 10 μg ofpurified RNA was mixed with 150 ng Random Hexamer primers (Invitrogen,Carlsbad, Calif., USA, catalog number: 48190-011) and 500 μM dNTPs in atotal volume of 156 μl. The mixture was incubated for 5 min at 65° C.and then quickly chilled on ice. Thereafter, 50 μl of 5× SuperscriptIIfirst strand buffer (Invitrogen, catalog number: 18064-014, part number:Y00146), 24 μl 0.1 M DTT and 400 units RNasin (Promega, Milwaukee, Wis.,U.S.A., catalog number: N2511) were added, and the mixture was incubatedfor 10 min at 25° C., followed by further incubation at 42° C. for 2min. Then, 10 μl (2000 units) of SuperscriptII (Invitrogen, catalognumber: 18064-014) was added and the reaction (final volume of 2500) wasincubated for 50 min at 42° C. and then inactivated at 70° C. for 15min. The resulting cDNA was diluted 1:20 in TE buffer (10 mM Tris, 1 mMEDTA pH 8).

2. PCR was done using Platinum PFX™ (Invitrogen., Carlsbad, Calif., USA,catalog number: 1178-021) under the following conditions: 5 μl PlatinumPFX 10× buffer; 5 μl-cDNA from the above; 2 μl-10 mM dNTPs (2.5 mM ofeach nucleotide); 0.5 μl-Platinum PFX enzyme; 37 μl-H2O; and 1.5 μl-ofeach primer (15 μM) in a total reaction volume of 50 μl; with a reactionprogram of 5 minutes in 95° C.; 35 cycles of: 30 seconds at 94° C., 30seconds at 55° C., 50 seconds at 68° C.; then 10 minutes at 68° C.Primers which were used include gene specific sequences corresponding tothe desired coordinates of the protein and restriction enzyme sites andKozak sequence, as listed in table 136, below. Bold letters in Table 136represent the specific gene sequence while the restriction siteextensions utilized for cloning purposes are in Italic and kozaksequences are underlined.

Table 136 demonstrates the cloning steps of ORF targets. For example,FXYD3_T25_P14 and VSIG1_T6_P5 were cloned by PCR amplification of twooverlapping fragments of the full length at step 1, followed byadditional PCR at step 2 using both PCR fragments from step 1 as atemplate for generating the full length. VSIG1_T5_P4 was cloned usingboth PCR fragments generated at step 1 for digestion and directligation, AI216611_T1_P1 was cloned by performing nested PCR on the PCRproduct generated from step 1. 5 μl of products No. 1, 4, 5, 8, 9, 10,11, 12, 15, 16 and 17 (Table 136), were loaded onto a 1% agarose gelstained with ethidium bromide, electrophoresed in 1×TBE solution at100V, and visualized with UV light. After verification of expected sizeband, remaining PCR product was processed for DNA purification usingQiaquick PCR purification kit (Qiagen™, Valencia, Calif., U.S.A.,catalog number 28106). The extracted PCR products were digested with theappropriate restriction enzymes (New England Biolabs, Beverly, Mass.,U.S.A.), as listed in table 136. After digestion, DNAs were loaded ontoa 1% agarose gel as described above. The expected band size was excisedand extracted from the gel using QiaQuick™ Gel Extraction kit (Qiagen,catalog number: 28707).

The digested targets' ORF DNAs were ligated to EGFP_pIRESpuro3 vectorusing the LigaFast™ Rapid DNA Ligation System (Promega, catalog number:M8221). The resulting DNAs were transformed into competent E. Colibacteria DH5α (RBC Bioscience, Taipei, Taiwan, catalog number: RH816)according to manufacturer's instructions, then plated on LB-ampicillinagar plates for selection of recombinant plasmids, and incubatedovernight at 37° C.

The following day, a number of colonies from each transformation thatgrew on the selective plates were taken for further analysis bystreak-plating on another selective plate and by PCR using GoTaqReadyMix (Promega, catalog number: M7122). Screening positive clones wasperformed by PCR using pIRESpuro3 vector specific primer and genespecific primer (data not shown). After completion of all PCR cycles,half of the reaction was analyzed using 1% agarose gel as describedabove. After verification of expected size band, 2 positive coloniesfrom each ligation reactions were grown in 5 ml Terrific Brothsupplemented with 100 μg/ml ampicillin, with shaking overnight at 37° C.Plasmid DNA was isolated from bacterial cultures using Qiaprep™ SpinMiniprep Kit (Qiagen, catalog number: 27106). Accurate cloning wasverified by sequencing the inserts (Weizmann Institute, Rehovot,Israel). Upon verification of an error-free colony (i.e. no mutationswithin the ORF), recombinant plasmids were processed for furtheranalysis.

The DNA sequences of the resulting VSIG1, ILDR1, LOC253012, AI216611,C1ORF32 or FXYD3 full length_fused to EGFP are shown in FIGS. 56A-J. InFIGS. 56A-J gene specific sequence correspond to the target's fulllength sequence is marked in bold faced, EGFP sequence is unbold Italicand known SNPs/silence mutations are underlined. FIG. 56A presents theDNA sequence of FXYD3_T0_P0_EGFP (996 bp) (SEQ ID NO:77); FIG. 56Bpresents the DNA sequence of FXYD3_T25_P14_EGFP (1083 bp) (SEQ IDNO:78); FIG. 56C presents the DNA sequence of AI216611_T0_P0_EGFP (1371bp) (SEQ ID NO:79); FIG. 56D presents the DNA sequence ofAI216611_T1_P1_EGFP (1332 bp) (SEQ ID NO:80); FIG. 56E presents the DNAsequence of C1ORF32_T8_P8_EGFP (1533 bp) (SEQ ID NO:81); FIG. 56Fpresents the DNA sequence of LOC253012_T4_P5_EGFP (2085 bp) (SEQ IDNO:82); FIG. 56G presents the DNA sequence of ILDR1_T0_P3_EGFP DNAsequence (2373 bp) (SEQ ID NO:83); FIG. 56H presents the DNA sequence ofILDR1_T2_P5_EGFP (2241 bp) (SEQ ID NO:84); FIG. 56I presents the DNAsequence of VSIG1_T6_P5_EGFP (2082 bp) (SEQ ID NO:85); FIG. 56J presentsthe DNA sequence of VSIG1_T5_P4_EGFP DNA (2004 bp) (SEQ ID NO:86).

The amino acid sequences of the resulting VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 or FXYD3 full length fused to EGFP are shown in FIG.57A-J; gene specific sequence correspond to the full length sequence ofthe protein is marked in bold faced, EGFP sequence is unbold Italic andamino acids modified due to known SNPs are underlined. FIG. 57A presentsthe amino acid sequence of FXYD3_P0_EGFP protein (331aa) (SEQ ID NO:87);FIG. 57B presents the amino acid sequence of FXYD3_P14_EGFP protein(360aa) (SEQ ID NO:88); FIG. 57C presents the amino acid sequence ofAI216611_P0_EGFP protein (456aa) (SEQ ID NO:89); FIG. 57D presents theamino acid sequence of AI216611_P1_EGFP protein (443aa) (SEQ ID NO:90);FIG. 57E presents the amino acid sequence of C1ORF32_P8_EGFP protein(510aa) (SEQ ID NO:91); FIG. 57F presents the amino acid sequence ofLOC253012_P5_EGFP protein (694aa) (SEQ ID NO:92); FIG. 57G presents theamino acid sequence of ILDR1_P3_EGFP protein (790aa) (SEQ ID NO:93);FIG. 57H presents the amino acid sequence of ILDR1_P5_EGFP protein(746aa) (SEQ ID NO:94); FIG. 57I presents the amino acid sequence ofVSIG1_P5_EGFP protein (693aa) (SEQ ID NO:95); FIG. 57J presents theamino acid sequence of VSIG1_P4_EGFP protein (667aa) (SEQ ID NO:96).

TABLE 136 full length cloning details

ull PCR Primer Restric- CGEN Target length product DNA Primer Primerorienta- tion ID name (aa) No template ID sequence tion site CGEN789FXYD3_

7 1 ovary 100-813 CTAGCTA GC For NheI T0_P0 30, 39, 59 (SEQ ID CACCATGCAGAA cDNA NO: 254) GGTGACCCTG (Table 4) 100-814 CGCGACCGG Rev AgeI(SEQ ID TCCGCTTTGGGC NO: 255) TGAGCCTGG FXYD3_ 16 2 lung 100-813 CTAGCTAGC For NheI T25_P14 1, 19, 20, (SEQ ID CACC ATGCAGAA 37, 42 NO: 254)GGTGACCCTG cDNA (Table 3) 100-843 CCTGTGTCC Rev (SEQ ID TCTTCATCTGTCNO: 256) 3 lung 100-842 GACAGATGA For 1, 19, 20, (SEQ ID AGAGGACACAGG37, 42 NO: 257) cDNA (Table 3) 100-814 CGCGACCGG Rev AgeI (SEQ IDTCCGCTTTGGGC NO: 255) TGAGCCTGG 4 PCR 100-813 CTAGCTA GC For NheIproducts (SEQ ID CACC ATGCAGAA No 2 + 3 NO: 254) GGTGACCCTG above100-814 CGCGACCGG Rev AgeI (SEQ ID TCCGCTTTGGGC NO: 255) TGAGCCTGGCGEN721 AI216611_

00 5 lung 49 100-740 CTAGCTA GC For NheI T0_P0 cDNA (SEQ ID CACCATGAGGCC (Table 3) NO: 258) TCTGCCCAGCG 100-741 CGCGAATTC Rev EcoRI(SEQ ID GACACTCAACAT NO: 259) CTTCCAGCTC AI216611_

99 6 lung 4 100-738 AAGGCTGCA For T1_P1 cDNA (SEQ ID TAGGAGCTG (Table 3)NO: 260) 100-919 CAATGAGTT Rev (SEQ ID GGAAATCAAGCC NO: 261) AC 7 PCR100-740 CTAGCTA GC For NheI product (SEQ ID CACC ATGAGGCC No 6 NO: 258)TCTGCCCAGCG above 100-919 CAATGAGTT Rev (SEQ ID GGAAATCAAGCCAC NO: 261)8 PCR 100-740 CTAGCTA GC For NheI product (SEQ ID CACC ATGAGGCC No 7NO: 258) TCTGCCCAGCG above 100-836 CGCGACCGG Rev AgeI (SEQ IDTCCAAACCACTC NO: 262) ATGGATTATCAC AAGGCCCAGGG GGTTACCTTTGA GTTTGTGTCTTCTC CGEN754 C1ORF32_

54 9 lung 100-746 CTAGCTA GC For NheI T8_P8 44, 45, 48 (SEQ ID CACCATGGATAG cDNA NO: 263) GGTCTTGCTGAG (Table 3) 100-694 CGCGAATTC RevEcoRI (SEQ ID GGGTAGAGAGGT NO: 264) AGACATTTC CGEN702 LOC253012_

50 10 IMAGE 100-765 GCGCTTCGA For BstBI T4_P5 clone (SEQ ID A GCCACCATGTG BC139906.1 NO: 265) GCTCAAGGTCTT CAC 100-766 CGCGACCGG Rev AgeI(SEQ ID TCCCTCTGGATG NO: 266) GTCTTGCTGCTG CGEN770 ILDR1_

46 12 ovary 100-780 CTAGCTA GC For NheI NheI T0_P3 19, 20, 27 (SEQ IDCACC ATGGCATG cDNA NO:267) GCCCAAACTGCC (Table 4) 100-781 CGCGACCGG RevAgeI (SEQ ID TCCAATGACCAC NO: 268) ACTCCTTCCACT A ILDRI_

02 12 ovary 100-780 CTAGCTA GC For NheI T2_P5 19, 20, 27 (SEQ ID CACCATGGCATG cDNA NO: 267) GCCCAAACTGCC (Table 4) 100-781 CGCGACCGG Rev AgeI(SEQ ID TCCAATGACCAC NO: 268) ACTCCTTCCACT A CGEN768 VSIG1_

49 13 lung 17 100-783 CTAGCTA GC For NheI T6_P5 cDNA (SEQ ID CACCATGGTGTT (Table 3) NO: 269) CGCATTTTGGAA G 100-838 CTGGAGTTC Rev (SEQ IDAGCCTGCTGTCCA NO: 270) TCAAGAG 14 lung 17 100-837 CTCTTGATG For cDNA(SEQ ID GACAGCAGGCTG (Table 3) NO: 271) AACTCCAG C 100-782 CGCGACCGG RevAgeI (SEQ ID TCCTGCCTTAAC NO: 272) CACTCCCTTTTC 15 PCR 100-783 CTAGCTAGC For NheI products (SEQ ID CACC ATGGTGTT No 13 + 14 NO: 269)CGCATTTTGGAA above G 100-782 CGCGACCGG Rev AgeI (SEQ ID TCCTGCCTTAACNO: 272) CACTCCCTTTTC VSIG1_

23 16 lung 4 100-783 CTAGCTA GC For NheI T5_P4 cDNA (SEQ ID CACCATGGTGTT (Table 3) NO: 269) CGCATTTTGGAA G 100-785 CCTC

Rev ScaI (SEQ ID

GAGGCACGAGC NO: 273) TGTG 17 lung 4 100-784 CCTC

For ScaI cDNA (SEQ ID

GAGGGTATGG (Table 3) NO: 274) 100-782 CGCGACCGG Rev AgeI (SEQ IDTCCTGCCTTAAC NO: 272) CACTCCCTTTTC

indicates data missing or illegible when filed

Example 9 Determining Cell Localization of VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 and FXYD3

In order to determine the cellular localization of the protein targets,they were cloned as EGFP (Enhanced Green Fluorescent Protein) fusionproteins. Proteins localization was observed upon transient transfection(Chen et al., Molecular vision 2002; 8; 372-388) using the confocalmicroscope. The cells were observed for the presence of fluorescentproducts 48 hours following transfection.

Determining cell localization of VSIG1, ILDR1, LOC253012, AI216611,C1ORF32 and FXYD3 was done by transient transfection of the recombinantORF-EGFP constructs which were described above.

The VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 and FXYD3-EGFP pIRESpuro3constructs were subsequently transiently transfected into HEK-293T cellsas follows:

HEK-293T (ATCC, CRL-11268) cells were plated on sterile glasscoverslips, 13 mm diameter (Marienfeld, catalog number: 01 115 30),which were placed in a 6 well plate, using 2 ml pre-warmed DMEM[Dulbecco's modified Eagle's Media, Biological Industries (Beit Ha'Emek,Israel), catalog number: 01-055-1A]+10% FBS (Fetal Bovin Serum)+4 mML-Glutamine. 500,000 cells per well were transfected with 2 μg of theDNA construct using 6 μl FuGENE 6 reagent (Roche, catalog number:11-814-443-001) diluted into 94 ul DMEM. The mixture was incubated atroom temperature for 15 minutes. The complex mixture was added dropwiseto the cells and swirled. Cells were placed in incubator maintained at37° C. with 5% CO₂ content.

48 hours post transient transfection the cells were further processedfor analysis in confocal microscopy. The cover slips were washed 3 timesin phosphate buffered saline (PBS) and fixed for 15 minutes with 3.7% or1% paraformaldehyde (PFA) (Sigma, catalog number: P-6148). After 2washes in PBS, the fixed coverslips were glued to a slide using mountingsolution (Sigma, catalog number: G0918) and cells were observed for thepresence of fluorescent product using confocal microscope. The resultsare presented in FIG. 58A-F.

FIG. 58A demonstrates that the AI216611_P0_EGFP (SEQ ID NO:89) andAI216611_P1_EGFP (SEQ ID NO:90) fused proteins localizes to cellmembrane upon expression in HEK 293T cells. The image was obtained usingthe 40× objective of the confocal microscope.

FIG. 58B demonstrates that the FXYD3_P0_EGFP (SEQ ID NO:87) andFXYD3_P14_EGFP (SEQ ID NO:88) fused proteins localizes to cell membraneupon expression in HEK 293T cells. The image was obtained using the 40×objective of the confocal microscope.

FIG. 58C demonstrates that the C1ORF32_P8_EGFP (SEQ ID NO:91) fusedprotein localizes to cell membrane; endoplasmatic reticulum (ER)membrane and to cell junctions upon expression in HEK 293T cells. Theimage was obtained using the 40× objective of the confocal microscope.

FIG. 58D demonstrates that the LOC253012_P5_EGFP (SEQ ID NO:92) fusedprotein localizes to cell membrane and endoplasmatic reticulum (ER)membrane upon expression in HEK 293T cells. The image was obtained usingthe 40× objective of the confocal microscope.

FIG. 58E demonstrates that the VSIG1_P5_EGFP (SEQ ID NO:95) andVSIG1-_P4_EGFP (SEQ ID NO:96) fused proteins localizes to nuclear cellmembrane and endoplasmatic reticulum membrane upon expression in HEK293T cells. The image was obtained using the 40× objective of theconfocal microscope.

FIG. 58F demonstrates that the ILDR1_P3_EGFP (SEQ ID NO:93) andILDR1_P5_EGFP (SEQ ID NO:94) fused proteins localizes to cell membraneand endoplasmatic reticulum membrane upon expression in HEK 293T cells.The image was obtained using the 40× objective of the confocalmicroscope.

Example 10 Cloning and Expression of VSIG1, ILDR1, LOC253012, AI216611,C1ORF32 and FXYD3 Extra Cellular Domain (ECD) Fused to Mouse Fc

The purpose of this analysis was to clone the VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 and FXYD3 ECDs fused via its corresponding C′ terminusto mouse Fc (mFc), and to express the fused ECDs in HEK293T cells(ATCC-CRL-11268), in order to be further used for antibody production aswell as for functional assessment of VSIG1, ILDR1, LOC253012, AI216611,C1ORF32 and FXYD3 ECDs.

The coordinates of the cloned ECD are described in table 137:

TABLE 137 Recombinant Full ECD protein length Coordinates CGEN ID nameTranscript No. Protein No. (aa) (aa) SEQ ID CGEN789 FXYD3 T25 P14 1161-63 SEQ ID No.- 297 (SEQ ID NO: 66) (SEQ ID NO: 72) CGEN721 AI216611 T0P0 200  1-145 SEQ ID No.- 298 (SEQ ID NO: 41) (SEQ ID NO: 43) CGEN754C1ORF32 T8 P8 254  1-184 SEQ ID No.- 299 (SEQ ID NO: 45) (SEQ ID NO: 48)CGEN702 LOC253012 T4 P5 450  1-335 SEQ ID No.- 300 (SEQ ID NO: 26) (SEQID NO: 36) CGEN770 ILDR1 T0 P3 546 51-160 SEQ ID No.- 301 (SEQ ID NO:17) (SEQ ID NO: 22) CGEN768 VSIG1 T6 P5 449 26-293 SEQ ID No.- 302 (SEQID NO: 7) (SEQ ID NO: 13)

The cloning of the fusion proteins (ECD_mFc) was done in two steps:

1. Cloning of ECD to pIRESpuro3.

2. Subcloning of the mouse Fc IgG2a in frame to the C′ terminus of theECD previously cloned into pIRESpuro3, from step1.

The cloning of ECD to pIRESpuro3 was carried out as follows:

Cloning of the ECD for each one of the VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 and FXYD3 was done by PCR delimit partial amino acidssequence of its ECD as described in table 137, using its full lengthsequence as a template, and primers as listed in table 138.

TABLE 138 ECD cloning details Primer restric- CGEN candidate primerorienta- tion ID name ID primer sequence tion site CGEN789 FXYD3 100-813CTAGCTA GCCACC ATGCAGAAGG For NheI SEQ ID TGACCCTG NO: 254 100-852CGCGGATCC CCAGTCTAGGAGGA Rev BamHI SEQ ID AAGTGG NO: 275 CGEN721AI216611 100-740 CTAGCTA GCCACC ATGAGGCCTC For NheI SEQ ID TGCCCAGCGNO: 258 100-850 CGCGGATCC GTCTTCATAGAGGA Rev BamHI SEQ ID TCTCAG BamHINO: 276 CGEN754 C1ORF32 100-746 CTAGCTA GCCACC ATGGATAGGG For NheISEQ ID TCTTGCTGAG NO: 263 100-851 CGCGGATCC CATAATCTCCACAG Rev BamHISEQ ID CAAAAC NO: 277 CGEN702 LOC253012 100-789 AACCGGT GCCACCATGTGGCTCA For AgeI SEQ ID AGGTCTTCAC NO: 278 100-854 CGCGGATCCTTTTCCTTTCTGTGC Rev BamHI SEQ ID AAGCT NO: 279 CGEN770 ILDRI 100-873GCGTTCGAA GCCCAGCTCCAGGA For BstBI SEQ ID CGTGGTG NO: 280 100-853CGCGGATCC TTCCTTATCGGGGT Rev BamHI SEQ ID CTCCTG NO: 281 CGEN768 VSIG1100-867 GCGCTTCGAA ATCCCAGACGGTT For BstBI SEQ ID TCGTG NO: 282 100-855CGCGGATCC TGGATGTGAAGAAG Rev BamHI SEQ ID TGAGAT NO: 283

In Table 138, above the bold letters represent the gene specificsequence while the restriction site extensions utilized for cloningpurposes are Italic and Kozak sequence is underlined.

The PCR products were purified and digested with the appropriaterestriction enzymes as describe in table 138. PCR products for FXYD3,AI216611, C1ORF32 and LOC253012 were ligated into pIRESpuro3, while PCRproducts for VSIG1 and ILDR1 were ligated into IL6sp_pIRESpuro3 in orderto increase their secretion. The ligation mixture was transformed intoDH5a competent cells. Positive transformants were screened and verifiedby DNA sequencing.

Cloning of ECD-mFc pIRESpuro3

Mouse Fc (IgG2a) (Accession-CAA49868 aa 237-469) protein sequencefollowed by TEV cleavage site sequence was codon optimized to boostprotein expression in mammalian system. The optimized sequence wassynthesized by GeneArt (Germany) with flanking BamHI restriction site atthe N′ terminus and NotI restriction site at the C′ terminus. The DNAfragment was digested with BamHI/NotI and ligated in frame intoECD_pIRESpuro3 constructs previously digested with the same enzymes togive ECD_mFc_pIRESpuro3. The ligation mixture was transformed into DH5acompetent cells. Positive transformants were screened and verified byDNA sequencing.

The nucleotide sequences of the resulting ECD_mFc ORFs are shown in FIG.59A-F: gene specific sequence correspond to the ECD sequence is markedin bold faced, TEV cleavage site sequence is underlined, mFc sequence isunbold Italic and IL6sp sequence is bold Italic. FIG. 59A shows theFXYD3_T25_P14_ECD-_mFc DNA sequence (924 bp) (SEQ ID NO:97); FIG. 59Bshows the AI216611_T0_P0_ECD_mFc DNA sequence (1170 bp) (SEQ ID NO:98),FIG. 59C shows the C1ORF32_T8_P8_ECD_mFc DNA sequence (1287 bp) (SEQ IDNO:99); FIG. 59D shows the LOC253012_T4_P5_ECD_mFc DNA sequence (1740bp) (SEQ ID NO:100), FIG. 59E shows the ILDR1_T0_P3_ECD_mFc DNA sequence(1167 bp) (SEQ ID NO:101), and FIG. 59F shows the VSIG1_T6_P5_ECD_mFcDNA sequence (1641 bp) (SEQ ID NO:102).

The sequence of the resulting ECD_mFc fusion proteins are shown in FIG.60A-60F; gene specific sequence correspond to the ECD sequence is markedin bold faced, TEV cleavage site sequence is underlined, mFc sequence isunbold Italic and IL6sp sequence is bold Italic. FIG. 60A shows theFXYD3_T25_P14_ECD-_mFc amino acid sequence (307aa) (SEQ ID NO:103); FIG.60B shows the AI216611_T0_P0_ECD_mFc amino acid sequence (389aa) (SEQ IDNO:104), FIG. 60C shows the C1ORF32_T8_P8_ECD_mFc amino acid sequence(428aa) (SEQ ID NO:105); FIG. 60D shows the LOC253012_T4_P5_ECD_mFcamino acid sequence (579aa) (SEQ ID NO:106), FIG. 60E shows theILDR1_T0_P3_ECD_mFc amino acid sequence (388aa) (SEQ ID NO:107), andFIG. 60F shows the VSIG1_T6_P5_ECD_mFc amino acid sequence (546aa) (SEQID NO:108).

To generate ECD-mFc expressing cells, HEK-293T cells were transfectedwith the above described constructs corresponding to VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 and FXYD3 extra cellular domain fused tomouse Fc. Stable pools were generated as follows 48 hrs posttransfection, the cells were trypsinized and transferred to T75 flaskcontaining selection medium (DMEM 10% FCS and 5 μg/ml puromycin) forobtaining stable pool. Media was changed every 3 to 4 days untilcolonies formation.

To verify the identity of cells, genomic PCR was performed, indicatingthe correct sequences integrated into the cell genome (data not shown).

Cell-deprived medium was collected and purified by Protein A-Sepharosebeads (Amersham catalog number 17-5280-04) as follows: 1 ml ofcell-deprived medium was incubated with 50 μl Protein A sepharose beadsfor 45 minutes at room temperature. At the end of incubation timeproteins were eluted from the beads pellet with 50 μl sample buffercontaining 100 mM Citrate Phosphate pH 3.5 and 10 mM DTT. The sampleswere boiled for 3 minutes and 25 μl were loaded on 12% NuPAGE Bis Trisgel (Invitrogen, catalog number NPO342). The proteins were transferredto a nitrocellulose membrane and blocked with 10% low fat milk in PBST(PBS supplemented with 0.05% tween-20). The membrane was then blottedfor 1 hour with Goat anti mouse IgG Fc fragment HRP (Jackson, catalognumber 115-035-206.) (1:40,000 in blocking solution) at roomtemperature. Following incubation with ECL solution (AmershamBiosciences, Catalog No. RPN2209), the membrane was exposed to film.

FIG. 61 shows the results of a western blot on expressed FXYD3_ECD_mFc(SEQ ID NO:103), AI216611 ECD_mFc (SEQ ID NO:104), C1ORF32_ECD_mFc (SEQID NO:105), LOC253012_ECD_mFc (SEQ ID NO:106), ILDR1_ECD_mFc (SEQ IDNO:107), VSIG1_ECD_mFc (SEQ ID NO:108) according to the presentinvention.

The lanes are as follows: lane 1 Molecular weight markers (Amersham,full range ranbow, catalog number RPN800); lane 2—LOC253012_ECD_mFc (SEQID NO:106); lane 3—FXYD3_ECD_mFc (SEQ ID NO:103); lane 4—AI216611ECD_mFc (SEQ ID NO:104); lane 5—C1ORF32_ECD_mFc (SEQ ID NO:105); lane6—ILDR1_ECD_mFc (SEQ ID NO:107); lane 7—VSIG1_ECD_mFc (SEQ ID NO:108).

Example 11 Protein Production of VSIG1, ILDR1, LOC253012, AI216611,C1ORF32 and FXYD3 Extra Cellular Domain (ECD) Fused to Mouse Fc

To produce VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 and FXYD3 ECDfused to mouse Fc, pool of transfected HEK293T cells stably transfectedwith the corresponding constructs described herein above, were used. Thetransfected cells, usually maintained in 10% serum supplemented medium,were transferred into serum free medium (EX-CELL293, SAFC) supplementedwith 4 mM glutamine and selection antibiotics (5 ug/ml puromycin), andgrown in suspension in shake flasks at 37° C., with agitation. Theculture volume was increased by sequential dilutions until a productionphase of 3-4 days carried out in 2 L spinners flasks. Medium from thespinners was harvested, cleared from cells by centrifugation, filteredthrough a 0.22 μm filter and kept at −20° C.

The VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 and FXYD3 ECD fused tomouse Fc were purified using nProtein A-affinity chromatography asdescribed below.

Harvests were concentrated approximately 10 fold using PALLultrafiltration system on two 10 kD cassettes. The concentrate was thenadjusted to pH 7.5, by the addition of 5M NaOH and filtrated through 0.2μm Stericup filter.

Purification process was carried out using AKTA Explorer (GEHealthcare). 2 ml of nProtein A Sepharose™, Fast Flow resin(cat#17-5280-02) were washed on Poly-prep chromatography column undervacuum with 10 column volumes (CV) of 70% ethanol, 10 CV WFI (SterileWater for Irrigation (TEVA)) followed by 10CV buffer A. 2 ml resin weretransferred into two 500 ml tubes (1 ml each) and the concentratedharvest was added. The tube was incubated overnight at 4° C. on a rollerto allow binding of the protein. Bound resin was then transferred andpacked under constant flow into XK16 column (GE Healthcare,cat#18-8773-01). The column was washed with 20 CV buffer A (100 Mm TrispH 7.4) and elution was carried out in one step using 100% buffer B(Citrate/Phosphate pH 3.0). The fractions were titrated with 12.5% (v/v)buffer C (2M Tris pH 8.5) to adjust the pH to −7.5 and pooled.

The final buffer was exchanged to DPBS (Dulbecco's Phosphate bufferssaline pH 7.4, w/o Ca, w/o Mg) pH 7.4 w/o Ca, w/o Mg using a 53 mlHiPrep™ (GE Healthcare, cat#17-5087-01) desalting column. The proteinwas filtered through 0.22 μm filter, aliquoted under sterile conditions,and stored at −800C.

The final protein concentration was determined by BCA total proteinassay and protein was analyzed by coomassie stained reducing SDS/PAGE(data not shown). Endotoxin level was determined by colorimetric LALassay (Limulus Amebocyte Lysate, QCL-1000, Cambrex). The identities ofthe specific proteins were verified by MS (at the Smoler ProteomicsCenter, Technion, Haifa, data not shown).

The resulted protein analyses are summarized in table 139.

TABLE 139 Concen- Puri- Endotox- tration ty ins Protein (mg/ml) (%)(EU/mg) C1ORF32-P8-ECD-mFc (SEQ 0.9 >90 1.04 ID NO: 105) IL6-VSIG1 P5ECD aa26-end 0.94 >95 0.95 mFc (SEQ ID NO: 108) FXYD3-T25P14-ECD-mFc1.1 >80 0.14 (SEQ ID NO: 103) AI216611-T0P0-mFc (SEQ ID 1.6 >94 0.72 NO:104) IL6 ILDR1 ECD aa50-160 mFc 1 85 <0.2 (SEQ ID NO: 107)LOC253012-P5-ECD-mFc 1 >95 1.45 (SEQ ID NO: 106)

Example 12 Binding of the ECDs Fc-Fused Proteins of the Invention toActivated T Cells

In order to examine of the ability of the VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 and FXYD3 Fc-fused ECDs described above to bind aputative counter-receptor on T cells, these Fc-fused ECDs were tested onresting or activated T cells. Purified T cells were activated with ConA(Sigma Aldrich, Cat #C5275), followed by incubation with the VSIG1,ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 Fc-fused ECDs and analyzedby flow cytometry.

T cells were purified from whole blood by negative selection usingRosetteSep™ Human T Cell Enrichment Cocktail (StemCell Technologies, CAT#15061). This resulted in a population of T (CD3+) cells with a purityof ˜90%. Purified T cells (1×105) were cultured for 48 hours in 100 ulof complete RPMI 1640 medium containing 10% FBS, either without anyactivation or activated with ConA (Concovalin A, 10 ug/ml, SigmaAldrich, Cat #C5275). Cultures were harvested and stained with the ECDsFc-fused proteins for 1 hour at 4° C. (VSIG1, ILDR1, LOC253012,AI216611, FXYD3 or C1ORF32 ECDs fused to mouse IgG2 Fc). The boundproteins were detected with FITC-conjugated F(ab)₂ goat anti-mouse Fcfor half an hour at 4° C. (Jackson ImmunoResearch Laboratories. CAT#115-096-071). Samples were analyzed using a FACSCalibur (BDImmunocytometry Systems) and CellQuest software.

FIGS. 62A-D present the binding of the ECDs Fc-fused proteins (VSIG1(SEQ ID NO:108), LOC253012 (SEQ ID NO:106), AI216611 (SEQ ID NO:104) orC1ORF32 (SEQ ID NO:105)) to resting T cells or T cells activated withCon A for different periods of time. Primary human T cells from threedifferent donors were cultured for a total of 48 hours in the absence ofstimulus (0 hrs) or in the presence of Con A, which was added to a finalconcentration of 10 μg/ml for the last 6, 18, 24 or 48 hours of culture(T cells from donor 5 were cultured with Con A for 0, 6, 18 and 24 hrs,while donors 6 & 7 were cultured for 0, 6, 24 and 48 hrs). Cells werethen harvested and incubated with 10 μg/ml of the indicated ECDsFc-fused proteins. FIG. 62A shows the binding results for Fc-fused VSIG1ECD; FIG. 62B shows the binding results for Fc-fused LOC253012; FIG. 62Cshows the binding results for Fc-fused C1ORF32 ECD; FIG. 62D shows thebinding results for Fc-fused AI216611 ECD and FIG. 62E shows the bindingresults for Fc-fused FXYD3 ECD. The percentage of positive cells wasdetermined as the difference between the positive cells with theindicated protein and the positive cells obtained with FITC-conjugatedF(ab)₂ goat anti-mouse Fc. FIG. 63 presents the dose response of thebinding of B7-like proteins to activated T cells. Purified T cells werecultured for 48 hours. Con A was added for the last 24 hours. Cells werethen harvested and stained with increasing concentrations (3, 6, 12, 25and 50 μg/ml) of Fc-fused VSIG1, LOC253012, C1ORF32, AI216611 orILDR1ECDs. As a negative controls, mouse IgG2a was used at the sameconcentrations.

The results presented in FIGS. 62A-D and 63 demonstrate binding of allthe ECDs Fc-fused proteins tested (VSIG1, ILDR1, LOC253012, AI216611 orC1ORF32 ECDs fused to mouse IgG2 Fc, SEQ ID NO:108, 107, 106, 104, or105, respectively), at binding levels above those of the negativecontrols: mouse IgG2a (R&D Systems, CAT #MAB003) as isotype control. Asubstantial binding was detected for Fc-fused VSIG1 ECD and forLOC253012 ECD-Fc to T cells stimulated with ConA. Fc-fused ECDs ofC1ORF32 and AI216611 showed a weaker binding to these cells, as can beseen from FIGS. 62A-D and 63. Each protein was found to bind a certainpercentage of activated T cells. The rating of binding levels was asfollows VSIG1>LOC253012>ILDR1=AI216611>C1ORF32. None of the proteinsbound resting T cells (i.e O hrs of ConA in FIGS. 62A-D).

Effect of the ECDs Fc-Fused Proteins of the Invention on T CellsActivation.

In order to test potential costimulatory or/and coinhibitory activity ofthe soluble proteins of the invention, VSIG1, ILDR1, LOC253012,AI216611, FXYD3 or C1ORF32 ECDs fused to mouse IgG2 Fc, SEQ ID NO:108,107, 106, 104, or 105, respectively, on T cells proliferation and IL-2secretion, human T cells were cultured in the presence of anti-CD3((clone OKT3, eBioscience, CAT #16-0037-85) and the B7-like proteins ofthe invention, described above. Recombinant human B7-1 protein (R&DSystems, CAT #140-B1) was used as a positive control for costimulatoryactivity. Recombinant mouse B7-H4 protein (R&D Systems, CAT #4206-B7)was used as positive control for coinhibitory activity.

Flat-bottom 96-well plates were first coated at 4° C. overnight with 3μg/ml of anti-CD3 mAb (clone OKT3) and subsequently coated with theindicated concentrations of human B7-1 (R&D, 3 μg/ml), mouse B7-H4 (R&D,10 μg/ml) or the ECDs Fc-fused proteins of the invention, VSIG1, ILDR1,LOC253012, AI216611, FXYD3 or C1ORF32 ECDs fused to mouse IgG2 Fc, for 4h at 37° C. Human T cells were purified from whole blood as describedabove, and were cultured in the pre-coated 96-well plates (1×105cells/well) in 250 μl of complete RPMI 1640 medium containing 10% FBSfor 48 hrs. Coated plates were washed with PBS three times beforeseeding of the cells. T cell proliferation was determined by BrdUincorporation by Cell proliferation ELISA, BrdU (colorimetric) (Roche).Cells were labeled with BrdU labeling reagent at a final concentrationof 100 μM for the last 18 hours. The plates were then centrifuged (at300 g, for 10 min), and supernatants were aspirated and stored at −20°C. for subsequent IL-2 determination using a Human IL-2 ELISA (Diaclone,CAT #850.010 096). BrdU incorporation was measured according toinstructions of the manufacturer of the Cell proliferation ELISA, BrdU(colorimetric) (Roche, CAT #11-647-229).

FIGS. 64A-B presents the effect of the ECDs Fc-fused proteins of theinvention on T cell proliferation or IL-2 secretion, upon activationwith anti-CD3 Ab. FIG. 64A shows the levels of BrdU incorporation. FIG.64B shows the levels of IL-2 secretion.

The results, presented in FIG. 64A-B, indicate that none of the ECD-Fcfused proteins VSIG1, ILDR1, LOC253012, AI216611, FXYD3 or C1ORF32 ECDsfused to mouse IgG2 Fc, showed costimulatory activity. The positivecontrol, B7-1, showed a strong costimulatory activity, as expected.Fc-fused ILDR1ECD and Fc-fused AI216611 ECD appear to have coinhibitoryactivity, since they inhibited cell proliferation similarly to B7-H4, incomparison to that obtained in the presence of the negative control:mouse IgG2a (FIG. 64A). However, no significant effect was observed onIL-2 secretion of any of the ECD-Fc fused proteins, VSIG1, ILDR1,LOC253012, AI216611, FXYD3 or C1ORF32 Fc-fused ECDs (FIG. 64B).

Example 13 Binding of the ECDs Fc-Fused Proteins of the Invention toLymphocytes and to CD4 Positive Cells

In order to further examine of the ability of the VSIG1, ILDR1,LOC253012, AI216611, FXYD3 and C1ORF32 Fc-fused ECDs to bind a putativecounter-receptor on T cells, these Fc-fused ECDs were tested first onlymphocytes. PBMCs were prepared from human peripheral blood, in FACSbuffer at 1×10e7/ml. Fc blocker (hIgG (16D10), lot#080706, 1.3 mg/ml) at30 ug/ml was added and cells were incubated with the blocker on ice for30 min. Fusion proteins were added at 1 ug/10e6 per stain on ice for 30min. 2nd Ab was added at 1 ug/100 ul/stain for 25-30 min(G@mIgG-Fc-FITC: Jackson Immunol Lab, 1 mg/ml, code#115-096-071,lot#71453, 1.0 mg/ml, used at 1 ug/stain). Cells were washed with thebuffer at each step outlined above. The binding was analyzed by flowcytometry.

FIG. 65 illustrates the binding of the ECDs Fc-fused of the VSIG1,ILDR1, LOC253012, AI216611, FXYD3 or C1ORF32 to lymphocytes. As can beseen from FIG. 65, C1ORF32, AI216611 and ILDR1 bind to a counterpartexpressed on lymphocytes.

Next, binding of the VSIG1, ILDR1, LOC253012, AI216611, FXYD3 andC1ORF32 Fc-fused ECDs to CD4+cells. Fc blocker (hIgG (16D10),lot#080706, 1.3 mg/ml) at 30 ug/ml was added and cells were incubatedwith the blocker on ice for 30 min. Fusion proteins were added at 1ug/10e6 per stain on ice for 30 min. Add 2nd Ab at 1 ug/100 ul/stain for25-30 min (G@mIgG-Fc-FITC: Jackson Immunol Lab, 1 mg/ml,code#115-096-071, lot#71453, 1.0 mg/ml, used at 1 ug/stain). @CD4(m@hCD4-APC: BD, cat3555349, lot#44331) was added 20 ul of each perstain, on ice for 30 min.

Cells were washed with the buffer at each step outlined above. Thebinding was analyzed by flow cytometry.

FIG. 66 illustrates the binding of the ECDs Fc-fused of ILDR1, C1ORF32and AI216611 to CD4+cells.

Example 14 Effect of the ECDs Fc-Fused Proteins of the Invention on TCell Activation

In order to test potential costimulatory or/and coinhibitory activity ofthe B7-like proteins of the invention, the affect of the VSIG1, ILDR1,LOC253012, AI216611 or C1ORF32 ECDs fused to mouse IgG2 Fc on T cellsproliferation was tested. T cells were purified from whole blood bypositive selection using CD3 microbeads (microbeads conjugated tomonoclonal anti-human CD3 antibodies (isotype: mouse IgG2a) (MACS WholeBlood CD3 Microbeads #130-090-874). Dynabeads are coated with CD3+/−B7with M-450 Epoxy Dynabeads (Invitrogen cat. No. 140.11). For activationof CD3 T cells, purified CD3 T cells are stimulated with theCD3+CD28coated beads at 1:1 or 1:05 ratio for various time points asneeded. The cells were seeded at 2×10e5 per well in presence or absenceof CD3+CD28 (2 ug/ml each)-coated beads and the cell proliferation wasmeasured after 72 hours by tritium—thymidine incorporation. The resultsare shown in FIG. 67. “CD3” in FIG. 67 mean CD3 only without thepresence of a costimulatory or coinhibitory molecule; “CD3+B7.2” meansCD3+a known B7 stimulatory control, B7.2; “CD3+B7H4” means CD3 and B7H4a known B7 inhibitory control; “CD3+B7H3” means CD3 and B7H3 a known B7stimulatory protein; “CD3+702” means CD3+LOC253012-ECD-Fc fused (SEQ IDNO:106); “CD3+721” means CD3+AI216611-ECD-Fc fused (SEQ ID NO:104);“CD3+754” means CD3+C1ORF32-ECD-Fc fused (SEQ ID NO:105); “CD3+768”means CD3+VSIG1-ECD-Fc fused (SEQ ID NO:108) “CD3+770” meansCD3+ILDR1_ECD-Fc fused (SEQ ID NO:107); “CD3+789” means CD3+FXYD3-ECD-Fcfused (SEQ ID NO:103).

As can be seen in FIG. 67, LOC253012-ECD-Fc, AI216611-ECD-Fc,VSIG1-ECD-Fc and FXYD3-ECD-Fc had an inhibitory effect on T cellscompared to CD3 alone in 3 different experiments (FIGS. 67 A, B, and C).

Example 15 Interaction of the ECDs-Fc Fused Proteins of the Inventionwith Resting B Cells, Activated B Cells, and B Cell Derived LymphomaCell Lines

Following demonstration of binding of the proteins of the invention tolymphocytes (Example 12 and 13, herein), the ability of the solubleproteins of the invention to bind to B cells was examined.

PBMCs were prepared from human peripheral blood, in FACS buffer at1×10e7/ml. Fc blocker (hIgG (16D10), lot#080706, 1.3 mg/ml) at 30 μg/mlwas added and cells were incubated with the blocker on ice for 30 min.Fusion proteins of invention ILDR1_ECD-Fc (SEQ ID NO:107),C1ORF32-ECD-Fc (SEQ ID NO:105), AI216611-ECD-Fc (SEQ ID NO:104),LOC253012-ECD-Fc (SEQ ID NO:106), FXYD3-ECD-Fc (SEQ ID NO:103), andVSIG1-ECD-Fc (SEQ ID NO:108) were added at 1 μg/10e6 per stain on icefor 30 minutes. 2nd Ab was added at 1 μg/100 ul/stain for 25-30 min(G@mIgG-Fc-FITC: Jackson Immunol Lab, 1 mg/ml, code#115-096-071,lot#71453, 1.0 mg/ml, used at 1 ug/stain). Cells were washed with thebuffer at each step outlined above. The binding was analyzed by flowcytometry. After that cells were stained with mouse @human IgM-PE (BDBioscience, CA, USA, cat#555783) which is specific for B cells. Thestained cells analyzed by flow cytometry. The @human IgM positive cellswere gated to analyze the binding of the fusion proteins of invention tothe B cells.

As shown in FIG. 68A, ILDR1_ECD-Fc and C1ORF32-ECD-Fc bound to B cellsof all 3 donors tested. AI216611-ECD-Fc exhibited binding to B cells in1 donor only.

In order to determine the existence of the counterpart on activated Bcells, PBMCs were activated with LPS for 72 hours with LPS. Thereafter,binding with the ECDs Fc-fused proteins of the invention ILDR1_ECD-Fc(SEQ ID NO:107), C1ORF32-ECD-Fc (SEQ ID NO:105), AI216611-ECD-Fc (SEQ IDNO:104), LOC253012-ECD-Fc (SEQ ID NO:106), FXYD3-ECD-Fc (SEQ ID NO:103),and VSIG1-ECD-Fc (SEQ ID NO:108) was done as described above, and cellswere stained with mouse @human CD86-Cy5PE (BD Bioscience, CA, USA, S,cat#555659) and mouse @human CD19-PE (BD Bioscience, CA, USA)antibodies.The activated B cells were defined as double positiveCD19+/CD86+population of cells.

As demonstrated in FIG. 68B, ILDR1_ECD-Fc (SEQ ID NO:107),C1ORF32-ECD-Fc (SEQ ID NO:105) and AI216611-ECD-Fc (SEQ ID NO:104)showed binding to activated B cells.

In order to determine the existence of the counterpart in B cellmalignancies, the binding of the ECDs Fc-fused proteins of inventionILDR1_ECD-Fc (SEQ ID NO:107), C1ORF32-ECD-Fc (SEQ ID NO:105),AI216611-ECD-Fc (SEQ ID NO:104), LOC253012-ECD-Fc (SEQ ID NO:106),FXYD3-ECD-Fc (SEQ ID NO:103), and VSIG1-ECD-Fc (SEQ ID NO:108) wereanalysed in B cell lymphoma cell lines. Raji (ATCC# CCL-86) and Daudi(ATCC# CCL-213) cells were purchased from ATCC and maintained inRPMI+10% FBS. The cells were stained with B7s protein or controls at 10μg/ml and thereafter with FITC-conjugated goat anti-mouse IgG Fc(Jackson Immunol Lab, NJ, USA, cat#115-096-071, lot#71453).

FIG. 68C illustrates the binding of the Fc-fused ECDs of the B7-likeproteins of the invention (ILDR1_ECD-Fc (SEQ ID NO:107), C1ORF32-ECD-Fc(SEQ ID NO:105), AI216611-ECD-Fc (SEQ ID NO:104), LOC253012-ECD-Fc (SEQID NO:106), FXYD3-ECD-Fc (SEQ ID NO:103), and VSIG1-ECD-Fc (SEQ IDNO:108)) to the B cell lymphoma cell lines. ILDR1_ECD-Fc (SEQ ID NO:107)showed a clear binding the both B cell lymphoma cell lines.

Example 16 Interaction of the ECD-Fc Fused Proteins of the Inventionwith Known B7

The interaction of AI216611 proteins of the invention with various knownligands of the B7 family was analyzed. Since AI216611 was predicted as apresumed CD28 receptor it was hypothesized to bind to a known B7 ligand,B7H4, which is considered orphan (its counterpart receptor has not yetbeen recognized).

The analysis of the interaction between B7H4-Ig (R&D Systems, Inc.cat.#4206-B7) and Fc fused AI216611 ECD (SEQ ID NO:104) was conductedusing the BIAcore 3000 system (Uppsala, Sweden) (Pharmacia Biosensor,Uppsala, Sweden) that employs surface Plasmon resonance for directlymeasuring intermolecular interactions. Fc fused AI216611 ECD (SEQ IDNO:104) (400-500 resonance units (RU)) was immobilized directly to thesensor CM5 chip. Solution containing two different concentrations ofB7H4-Ig (5 and 10 micro molar) was injected. As control, the solutionswere also injected onto an empty flow cell with no ligand immobilized.

Data was analyzed using BIAevaluation software (GraphPad Software Inc.,San Diego Calif.). A zero baseline level was obtained by subtracting thebackground responses from injection of the analytes through a controlflow cell with no ligand immobilized.

As can be seen from FIG. 69, a slight interaction between Fc fusedAI216611 ECD (SEQ ID NO:104) and B7H4 was found in 5 and 10 μM ofAI216611.

Example 17 Development of Mouse Monoclonal Anti-VSIG1, Anti-ILDR1,Anti-LOC253012, Anti-AI216611, Anti-C1ORF32 and Anti-FXYD3 Antibodies

In order to test the expression of B7-Like proteins in different cancertissues by immunohistochemistry, monoclonal mouse antibodies specificfor Fc-fused ECDs of the proteins of invention were developed.

Development of Mouse Monoclonal Antibodies:

Four groups of the Balb/c mice (3 mice per group) were immunized with 4Fc-fused ECDs proteins of the invention: VSIG1 (SEQ ID NO:108),LOC253012 (SEQ ID NO:106), C1ORF32 (SEQ ID NO:105) and FXYD3(SEQ IDNO:103). The immunizations were performed 8 times at one week intervalsin multiple sites, subcutaneous and intraperitoneal. Mice were bled tendays following the 4th and 8th immunizations. Serum was screened forantibody titer using a Direct ELISA protocol described below.

ELISA plates were coated with 50 μl/well of 2.5 μg/mL Fc-fused proteins(VSIG1, LOC253012, C1ORF32, FXYD3 ECDs fused to mouse IgG2 Fc, SEQ IDNOs: 108, 106, 105, 103, respectively) diluted in DPBS for 1 hour atroom temperature (RT). Human IgG fused to mouse Fc region was used as anegative control. After that, plates were blocked with 300 μl/well of 1%BSA/DPBS for 15 min at RT. Following the blocking step, serially dilutedsera from immunized mice and irrelevant mouse IgG were transferred tothe blocked ELISA plates and incubated for 1 hour at RT. Afterwards,plates were washed 3 times with 300 μl/well washing buffer (DPBS with0.05% Tween 20, pH 7.2-7.4). For detection, plates were incubated for 1hour at RT with 50 μl/well of Goat anti-Mouse Kappa Light Chain Antibodyat 1:1000 dilution followed by an extensive wash (6 times with 300μl/well of washing buffer) and incubation with the substrate. Thesubstrate, 2,2′-Azino-bis-(3-ethylbenzthiazoline-6-sulfonic acid (ABTS),at 100 μL/well was added and incubated for about 5 min at RT beforeplates were read at 414 nm using a Molecular Devices SPECTRAmax 340 PCplate reader and SOFTmax PRO software.

Serum antibody titer was defined as the dilution of serum that producesa signal that was twice that of the background.

Results of the ELISA test of the immunized sera after 4 immunizationsare summarized in the Table 140. Data show that after 4 immunizations, 2mice groups (immunized with LOC253012 and VSIG1 Fc fused proteins ECDs)developed antibody titers sufficient for hybridoma production.

The mice that showed highest antibody serum titers, were selected forhybridoma production. The splenocytes were fused with mouse myeloma cellline Ag8.653. The supernatant of the hybridoma clones were tested bydirect ELISA (as described above) using plates coated with relevant andirrelevant coatings. The results are summarized in Table 141A and Table141B.

The results demonstrate that production of hybridoma cell lines resultedin 14 clones specifically recognizing LOC253012 (Table 141A, bold) and14 clones specifically recognizing VSIG1 (Table 141B, bold).

For the rest of the proteins, four additional immunizations wereperformed in order to facilitate the serum antibody titers developmentfor the rest of the proteins. The sera titers after the 8th immunizationwere tested by direct ELISA. Results are summarized in Table 142. Theresults demonstrate that after 8 immunizations the mice immunized withFXYD-Fc fused ECD (SEQ ID NO:103) and C1ORF32-Fc fused ECD (SEQ IDNO:103) developed sufficient antibody titers for hybridoma production.In the next step, the best responders will be selected for hybridomaproduction and monoclonal antibody manufacturing.

Mouse monoclonal anti-ILDR1 and anti-AI216611 antibodies are developedsimilarly.

The Monoclonal Antibodies for each of the antigens of the invention(VSIG1, LOC253012, C1ORF32, FXYD3, AI216611 and ILDR1, SEQ ID NOs: 108,106, 105, 103, 104 and 107, respectively) are used forImmunohistochemistry analysis in order to verify the expression profileof each of these putative proteins in cancer and healthy tissues.

TABLE 140 Antibody sera titers of the immunized mice after 4immunizations. ELISA Plates Coatings. human Immunogen Mouse # C1ORF32FXYD3 LOC253012 VSIG1 IgG-moFc C1ORF32 167229 378 212 <50 181 152 167230612 319 <50 383 159 167231 599 445 276 934 398 FXYD3 167232 1,409 2,532962 2,229 1,433 167233 1,379 2,320 695 2,777 1,211 167234 1,585 4,604615 3,625 1,751 LOC253012 167223 <50 51 18,869 <50 68 167224 <50 939,939 73 156 167225 93 560 3,025 268 116 VSIG1 167226 <50 <50 <50 10,653<50 167227 158 603 <50 18,085 58 167228 412 751 58 93,059 83

TABLE 141A Post fusional clones resulted from mouse #167223, immunizedwith LOC253012 ELISA Plates Coatings C1ORF32 FXYD3 LOC253012 VSIG1 moFCClone ID OD at 441 nm 2G2 0.188 0.214 2.296 0.216 0.278 3A8 2.053 2.4501.926 1.787 0.326 4A8 0.201 0.225 2.553 0.222 0.279 6B10 0.227 0.2062.335 0.227 0.293 8G10 0.476 0.346 1.562 0.267 0.487 8G11 0.192 0.2002.178 0.220 0.274 10A3 0.189 0.190 1.654 0.215 0.272 10F2 0.246 0.2471.720 0.242 0.350 12D5 0.198 0.190 1.619 0.224 0.291 13A4 0.252 0.2211.847 0.228 0.312 13F11 0.219 0.194 1.865 0.223 0.296 13H2 0.216 0.2291.404 0.255 0.300 14D11 0.199 0.230 2.183 0.225 0.294 16A10 1.285 2.1301.239 0.972 0.277 16C10 2.159 2.516 1.927 1.908 0.273 16F10 0.183 0.1790.235 0.203 0.271 17E5 0.188 0.193 1.943 0.214 0.270 18G4 0.202 0.2091.843 0.216 0.284

TABLE 141B Post fusional clones resulted from mouse #167228 immunizedwith VSIG1 ELISA Plates Coatings. C1ORF32 FXYD3 LOC253012 VSIG1 moFCClone ID OD at 441 nm 3F8 0.192 0.193 0.240 2.279 0.283 4D5 0.220 0.2060.251 2.174 0.299 4D6 0.206 0.207 0.227 2.808 0.279 5B6 0.197 0.1880.208 0.380 0.261 6G2 0.227 0.208 0.198 1.880 0.294 7C1 0.208 0.2120.209 2.392 0.278 7E3 2.284 2.804 1.495 2.278 0.398 7H3 0.207 0.2320.197 1.530 0.295 9F4 0.226 0.274 0.175 1.965 0.372 10B10 0.228 0.2740.219 0.407 0.330 11B2 0.214 0.247 0.207 2.733 0.316 11F3 2.638 3.0521.919 2.575 0.313 11G10 0.244 0.249 0.191 2.076 0.337 13A1 0.240 0.2390.195 2.469 0.316 13H5 2.782 2.886 1.963 2.305 0.293 14D8 0.218 0.2300.187 2.660 0.294 15E8 0.211 0.240 0.180 1.966 0.293 17E6 0.602 0.5470.271 0.341 0.662 19G6 0.439 0.490 0.245 1.279 0.318

TABLE 142 Antibody sera titers of the immunized mice after 8immunizations. ELISA Plates Coatings human Immunogen Mouse # C1ORF32FXYD3 LOC253012 VSIG1 IgG-moFc C1ORF32 167229 21,760 2,192 295 300 2,287167230 69,543 613 59 265 693 167231 23,100 1,952 756 1,645 2,487 FXYD3167232 3,237 58,240 1,564 2,622 6,515 167233 5,061 10,786 2,125 4,9597,664 167234 3,445 122,929 811 2,267 7,061 LOC253012 167223 Fused 1672242,641 18,011 328,050 5,491 260 167225 6,132 23,589 173,452 7,662 90VSIG1 167226 819 6,096 207 471,316 89 167227 39,852 102,238 18,463532,487 256 167228 Fused

Immunohistochemical Analysis

Immunohistochemistry enables the visualization (using light or confocalmicroscopy) of the tissue distribution of specific antigens (orepitopes). The process localizes protein targets of interest by applyingspecific monoclonal or polyclonal antibodies to tissue surfaces in aprocess called antibody incubation.

This method involves detection of a substrate in situ in fixed cells bysubstrate specific antibodies. The substrate specific antibodies may beenzyme linked or linked to fluorophores. Detection is carried out bymicroscopy and subjective evaluation. If enzyme linked antibodies areemployed, a colorimetric reaction may be required.

The immunohistochemical analysis performed for the antigens of theinvention (VSIG1, LOC253012, C1ORF32, FXYD3, AI216611 and ILDR1, SEQ IDNOs: 108, 106, 105, 103, 104 and 107, respectively) consist of twophases:

Phase I: Antibody calibration: A dilution series of each of theantibodies developed against the specific protein antigens is run usingselected formalin-fixed paraffin-embedded (FFPE) control tissues andcell lines. The best performing antibody is selected for Phase II.

Phase II: Protein distribution and localization analysis: Using theoptimal antibody concentration selected in Phase I, the distribution andlocalization of VSIG1, LOC253012, C1ORF32, FXYD3, AI216611 and ILDR1proteins is analyzed in Tissue Arrays consisting of cancer and healthytissues, looking for differential expression of the in some of thecancer samples, as compared with healthy samples.

Example 17 Development of Fully Human Anti-VSIG1, Anti-ILDR1,Anti-LOC253012, Anti-AI216611, Anti-C1ORF32 and Anti-FXYD3 Antibodies

Generation Of Human Monoclonal Antibodies Against VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 and FXYD3 Antigen

Fusion proteins composed of the extracellular domain of the VSIG1,ILDR1, LOC253012, AI216611, C1ORF32 and FXYD3 linked to a mouse IgG2 Fcpolypeptide are generated by standard recombinant methods and used asantigen for immunization.

Transgenic HuMab Mouse.

Fully human monoclonal antibodies to VSIG1, ILDR1, LOC253012, AI216611,C1ORF32 and FXYD3 are prepared using mice from the HCo7 strain of thetransgenic HuMab Mouse®, which expresses human antibody genes. In thismouse strain, the endogenous mouse kappa light chain gene has beenhomozygously disrupted as described in Chen et al. (1993) EMBO J.12:811-820 and the endogenous mouse heavy chain gene has beenhomozygously disrupted as described in Example 1 of PCT Publication WO01/09187. Furthermore, this mouse strain carries a human kappa lightchain transgene, KCo5, as described in Fishwild et al. (1996) NatureBiotechnology 14:845-851, and a human heavy chain transgene, HCo7, asdescribed in U.S. Pat. Nos. 5,545,806; 5,625,825; and 5,545,807.

HuMab Immunizations:

To generate fully human monoclonal antibodies to VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 and FXYD3, mice of the HCo7 HuMab Mouse®(strain can be immunized with purified recombinant VSIG1 fusion proteinderived from mammalian cells that are transfected with an expressionvector containing the gene encoding the fusion protein. Generalimmunization schemes for the HuMab Mouse® are described in Lonberg, N.et al (1994) Nature 368(6474): 856-859; Fishwild, D. et al. (1996)Nature Biotechnology 14: 845-851 and PCT Publication WO 98/24884. Themice are 6-16 weeks of age upon the first infusion of antigen. Apurified recombinant VSIG1 antigen preparation (5-50. mu.g, purifiedfrom transfected mammalian cells expressing VSIG1 fusion protein) isused to immunize the HuMab mice intraperitoneally.

Transgenic mice are immunized twice with antigen in complete Freund'sadjuvant or Ribi adjuvant IP, followed by 3-21 days IP (up to a total of11 immunizations) with the antigen in incomplete Freund's or Ribiadjuvant. The immune response is monitored by retroorbital bleeds. Theplasma is screened by ELISA (as described below), and mice withsufficient titers of anti-VSIG1 human immunoglobulin are used forfusions. Mice are boosted intravenously with antigen 3 days beforesacrifice and removal of the spleen.

Selection of HuMab Mice™ Producing Anti-VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 and FXYD3 Antibodies:

To select HuMab Mice™ producing antibodies that bind VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 sera from immunized mice is testedby a modified ELISA as originally described by Fishwild, D. et al.(1996). Briefly, microtiter plates are coated with purified recombinantVSIG1 fusion protein at 1-2. mu.g/ml in PBS, 50. mu.l/wells incubated 4degrees C. overnight then blocked with 200. mu.l/well of 5% BSA in PBS.Dilutions of plasma from VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 orFXYD3-immunized mice are added to each well and incubated for 1-2 hoursat ambient temperature. The plates are washed with PBS/Tween and thenincubated with a goat-anti-human kappa light chain polyclonal antibodyconjugated with alkaline phosphatase for 1 hour at room temperature.After washing, the plates are developed with pNPP substrate and analyzedby spectrophotometer at OD 415-650. Mice that developed the highesttiters of anti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611,anti-C1ORF32 or anti-FXYD3 antibodies are used for fusions. Fusions areperformed as described below and hybridoma supernatants are tested foranti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32 oranti-FXYD3 activity by ELISA.

Generation of Hybridomas Producing Human Monoclonal Antibodies to VSIG1,ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3

The mouse splenocytes, isolated from the HuMab mice, are fused with PEGto a mouse myeloma cell line based upon standard protocols. Theresulting hybridomas are then screened for the production ofantigen-specific antibodies. Single cell suspensions of spleniclymphocytes from immunized mice are fused to one-fourth the number ofP3X63 Ag8.6.53 (ATCC CRL 1580) nonsecreting mouse myeloma cells with 50%PEG (Sigma). Cells are plated at approximately 1×10−5/well in flatbottom microtiter plate, followed by about two week incubation inselective medium containing 10% fetal calf serum, supplemented withorigen (IGEN) in RPMI, L-glutamine, sodium pyruvate, HEPES, penicillin,streptomycin, gentamycin, 1×HAT, and beta-mercaptoethanol. After 1-2weeks, cells are cultured in medium in which the HAT is replaced withHT. Individual wells are then screened by ELISA (described above) forhuman anti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611,anti-C1ORF32 or anti-FXYD3 monoclonal IgG antibodies. Once extensivehybridoma growth occurred, medium is monitored usually after 10-14 days.The antibody secreting hybridomas are replated, screened again and, ifstill positive for human IgG, anti-VSIG1, anti-ILDR1, anti-LOC253012,anti-AI216611, anti-C1ORF32 or anti-FXYD3 monoclonal antibodies aresubcloned at least twice by limiting dilution. The stable subclones arethen cultured in vitro to generate small amounts of antibody in tissueculture medium for further characterization.

Hybridoma clones are selected for further analysis.

Structural Characterization of Desired Anti-VSIG1, Anti-ILDR1,Anti-LOC253012, Anti-AI216611, Anti-C1ORF32 or Anti-FXYD3 HumanMonoclonal Antibodies

The cDNA sequences encoding the heavy and light chain variable regionsof the obtained anti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611,anti-C1ORF32 or anti-FXYD3 monoclonal antibodies are obtained from theresultant hybridomas, respectively, using standard PCR techniques andare sequenced using standard DNA sequencing techniques.

The nucleotide and amino acid sequences of the heavy chain variableregion and of the light chain variable region are identified. Thesesequences may be compared to known human germline immunoglobulin lightand heavy chain sequences and the CDRs of each heavy and light of theobtained anti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611,anti-C1ORF32 or anti-FXYD3 sequences identified.

Characterization of Binding Specificity and Binding Kinetics ofAnti-VSIG1, Anti-ILDR1, Anti-LOC253012, Anti-AI216611, Anti-C1ORF32 orAnti-FXYD3 Human Monoclonal Antibodies

The binding affinity, binding kinetics, binding specificity, andcross-competition of anti-VSIG1, anti-ILDR1, anti-LOC253012,anti-AI216611, anti-C1ORF32 or anti-FXYD3 antibodies are examined byBiacore analysis. Also, binding specificity is examined by flowcytometry.

Binding Affinity and Kinetics

Anti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32 oranti-FXYD3 antibodies produced according to the invention arecharacterized for affinities and binding kinetics by Biacore analysis(Biacore AB, Uppsala, Sweden). Purified recombinant human VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 fusion protein is covalentlylinked to a CM5 chip (carboxy methyl dextran coated chip) via primaryamines, using standard amine coupling chemistry and kit provided byBiacore. Binding is measured by flowing the antibodies in HBS EP buffer(provided by BIAcore AB) at a concentration of 267 nM at a flow rate of50. mu.l/min. The antigen-association antibodies association kinetics isfollowed for 3 minutes and the dissociation kinetics is followed for 7minutes. The association and dissociation curves are fit to a 1:1Langmuir binding model using BIAevaluation software (Biacore AB). Tominimize the effects of avidity in the estimation of the bindingconstants, only the initial segment of data corresponding to associationand dissociation phases are used for fitting.

Epitope Mapping of Obtained Anti-VSIG1, Anti-ILDR1, Anti-LOC253012,Anti-AI216611, Anti-C1ORF32 or Anti-FXYD3 Antibodies

Biacore is used to determine epitope grouping of anti-VSIG1, anti-ILDR1,anti-LOC253012, anti-AI216611, anti-C1ORF32 or anti-FXYD3 HuMAbs.Obtained anti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611,anti-C1ORF32 or anti-FXYD3 antibodies are used to map their epitopes onthe VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 antigen,respectively. These different antibodies are coated on three differentsurfaces of the same chip to 8000 RUs each. Dilutions of each of themAbs are made, starting at 10 mu.g/mL and is incubated with Fc fusedVSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 (50 nM) for onehour. The incubated complex is injected over all the three surfaces (anda blank surface) at the same time for 1.5 minutes at a flow rate of 20.mu.L/min. Signal from each surface at end of 1.5 minutes, aftersubtraction of appropriate blanks, has been plotted againstconcentration of mAb in the complex. Upon analysis of the data, theanti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32 oranti-FXYD3 antibodies are categorized into different epitope groupsdepending on the epitope mapping results. The functional propertiesthereof are also compared.

Chinese hamster ovary (CHO) cell lines that express VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 protein at the cell surface aredeveloped and used to determine the specificity of the VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 HuMAbs by flow cytometry. CHOcells are transfected with expression plasmids containing full lengthcDNA encoding a transmembrane forms of VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 or FXYD3 antigen or a variant thereof. The transfectedproteins contained an epitope tag at the N-terminus are used fordetection by an antibody specific for the epitope. Binding of aanti-VSIG1, anti-ILDR1, anti-LOC253012, anti-AI216611, anti-C1ORF32 oranti-FXYD3 MAb is assessed by incubating the transfected cells with eachof the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 Abs at aconcentration of 10 mu.g/ml. The cells are washed and binding isdetected with a FITC-labeled anti-human IgG Ab. A murine anti-epitopetag Ab, followed by labeled anti-murine IgG, is used as the positivecontrol. Non-specific human and murine Abs are used as negativecontrols. The obtained data is used to assess the specificity of theHuMAbs for the VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3.antigen target.

These antibodies and other antibodies specific to VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 may be used in the afore-describedanti-VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 relatedtherapies such as treatment of cancers wherein VSIG1, ILDR1, LOC253012,AI216611, C1ORF32 or FXYD3 antigen is differentially expressed such aslung cancer, colon cancer and ovarian cancer and/or for modulating(enhancing or inhibiting) B7 immune co-stimulation involving the VSIG1,ILDR1, LOC253012, AI216611 or C1ORF32 antigen such as in the treatmentof cancers and autoimmune diseases wherein such antibodies will e.g.,prevent negative stimulation of T cell activity against desired targetcancer cells or prevent the positive stimulation of T cell activitythereby eliciting a desired anti-autoimmune effect.

The invention has been described and prophetic embodiments providedrelating to manufacture and selection of desired anti-VSIG1, ILDR1,LOC253012, AI216611, C1ORF32 or FXYD3 antibodies for use as therapeuticsand diagnostic methods wherein the disease or condition is associatedwith VSIG1, ILDR1, LOC253012, AI216611, C1ORF32 or FXYD3 antigen. Theinvention is now further described by the claims which follow.

1-221. (canceled)
 222. An isolated polypeptide comprising theextracellular domain of H19011_(—)1_P8 (SEQ ID NO:48) or H19011_(—)1_P9(SEQ ID NO:50).
 223. The polypeptide of claim 222 comprising a sequenceof amino acid residues having at least 95% sequence identity with aminoacid residues 21-186 of H19011_(—)1_P8 (SEQ ID NO:48), corresponding toamino acid sequence depicted in SEQ ID NO:147, or residues 21-169 ofH19011_(—)1_P9 (SEQ ID NO:50), corresponding to amino acid sequencedepicted in SEQ ID NO:148, or to amino acid sequence depicted in SEQ IDNO:299.
 224. The polypeptide of claim 223, wherein said polypeptide hasthe amino acid sequence of SEQ ID NO:147, SEQ ID NO:148 or SEQ IDNO:299.
 225. The polypeptide of claim 224, wherein said polypeptide hasthe amino acid sequence of SEQ ID NO:147.
 226. The polypeptide of claim224, wherein said polypeptide has the amino acid sequence of SEQ IDNO:148.
 227. The polypeptide of claim 224, wherein said polypeptide hasthe amino acid sequence of SEQ ID NO:299.
 228. The polypeptide of claim222, wherein said polypeptide blocks or inhibits the interaction ofH19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50), or afragment or variant thereof with a corresponding functional counterpart.229. A fusion protein comprising the polypeptide of claim 222 joined toa heterologous sequence.
 230. The fusion protein of claim 229, whereinthe heterologous sequence comprises at least a portion of animmunoglobulin molecule.
 231. The fusion protein of claim 229 having theamino acid sequence set forth in SEQ ID NO:
 105. 232. A nucleic acidsequence encoding the polypeptide of claim 222 or a fusion proteincomprising the polypeptide of claim 222 joined to a heterologoussequence.
 233. An expression vector containing at least one nucleic acidsequence according to claim
 232. 234. A recombinant cell comprising anexpression vector or a virus containing a nucleic acid sequenceaccording to claim 233, wherein the cell constitutively or induciblyexpresses the polypeptide encoded by the DNA segment.
 235. A method ofproducing a C1ORF32 ectodomain polypeptide, or fragment or fusionprotein thereof, comprising culturing the recombinant cell according toclaim 234, under conditions whereby the cell expresses the polypeptideencoded by the DNA segment or nucleic acid and recovering saidpolypeptide.
 236. The nucleic acid sequence according to claim 232,having the nucleic acid sequence set forth in SEQ ID NO:99.
 237. Thefusion protein of claim 229, which modulates lymphocyte activation invitro or in vivo.
 238. A pharmaceutical composition comprising at leastone of a polypeptide comprising the extracellular domain ofH19011_(—)1_P8 (SEQ ID NO:48) or H19011_(—)1_P9 (SEQ ID NO:50) or afusion thereof to a heterologous sequence; a nucleotide sequenceencoding the polypeptide comprising the extracellular domain ofH19011_(—)1_P8 (SEQ ID NO:48) or H19011_(—)1_P9 (SEQ ID NO:50) or afusion thereof to a heterologous sequence; an expression vectorcontaining; a host cell comprising an expression vector containing anucleic acid sequence encoding the polypeptide comprising theextracellular domain of H19011_(—)1_P8 (SEQ ID NO:48) or H19011_(—)1_P9(SEQ ID NO:50), and further comprising a pharmaceutically acceptablediluent or carrier.
 239. A method for treating or preventing immunerelated condition comprising administering to a subject in need thereofa pharmaceutical composition of claim
 238. 240. The method of claim 239,wherein the treatment is combined with a moiety useful for treatingimmune related condition.
 241. The method of claim 240, wherein themoiety is a cytokine antibody, cytokine receptor antibody, drug, oranother immunomodulatory agent.
 242. The method of claim 239 whereinsaid immune conditions are selected from autoimmune disease, transplantrejection, or graft versus host disease.
 243. The method of claim 242wherein the autoimmune disease is selected from a group consisting ofmultiple sclerosis; psoriasis; rheumatoid arthritis; systemic lupuserythematosus; Crohn's disease, ulcerative colitis, immune disordersassociated with graft transplantation rejection; benign lymphocyticangiitis, lupus erythematosus, Hashimoto's thyroiditis, primarymyxedema, Graves' disease, pernicious anemia, autoimmune atrophicgastritis, Addison's disease, insulin dependent diabetes mellitis, Goodpasture's syndrome, myasthenia gravis, pemphigus, sympatheticophthalmia, autoimmune uveitis, autoimmune hemolytic anemia, idiopathicthrombocytopenia, primary biliary cirrhosis, chronic action hepatitis,ulceratis colitis, Sjogren's syndrome, rheumatic disease, polymyositis,scleroderma, mixed connective tissue disease, inflammatory rheumatism,degenerative rheumatism, extra-articular rheumatism, collagen diseases,chronic polyarthritis, psoriasis arthropathica, ankylosing spondylitis,juvenile rheumatoid arthritis, periarthritis humeroscapularis,panarteriitis nodosa, progressive systemic scleroderma, arthritisuratica, dermatomyositis, muscular rheumatism, myositis, myogelosis andchondrocalcinosis.
 244. A monoclonal or polyclonal antibody or anantigen binding fragment thereof comprising an antigen binding site thatbinds specifically to any one of the C1ORF32 polypeptides comprised inH19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ ID NO:50); or toamino acid residues 21-186 of H19011_(—)1_P8 (SEQ ID NO:48),corresponding to amino acid sequence depicted in SEQ ID NO:147, or toamino acid residues 21-169 of H19011_(—)1_P9 (SEQ ID NO:50),corresponding to amino acid sequence depicted in SEQ ID NO:148, or toamino acid sequence depicted in SEQ ID NO:299, or a variant thereof thatis at least 95% identical thereto, or an epitope thereof.
 245. Theantibody or fragment of claim 244 wherein the antigen binding sitecomprises a conformational or linear epitope, and wherein the antigenbinding site contains from about 3-7 contiguous or non-contiguous aminoacids.
 246. The antibody or fragment according to claim 244, wherein theantibody is a fully human antibody, chimeric antibody, humanized orprimatized antibody.
 247. The antibody or fragment according to claim244, wherein the antibody is selected from the group consisting of Fab,Fab′, F(ab′)2, F(ab′), F(ab), Fv or scFv fragment and minimalrecognition unit.
 248. The antibody or fragment according to claim 244,wherein the antibody is coupled to a moiety selected from a drug, aradionuclide, a fluorophore, an enzyme, a toxin, a therapeutic agent, ora chemotherapeutic agent; and wherein the detectable marker is aradioisotope, a metal chelator, an enzyme, a fluorescent compound, abioluminescent compound or a chemiluminescent compound.
 249. Apharmaceutical composition that comprises an antibody or a fragmentaccording to claim
 244. 250. A method for potentiating a secondaryimmune response to an antigen in a patient, which method comprisesadministering effective amounts of at least one antibody or fragmentaccording to claim
 244. 251. The method of claim 250 wherein the antigenis a cancer antigen, a viral antigen or a bacterial antigen, and thepatient has received treatment with an anticancer vaccine or a viralvaccine.
 252. A method of treating or preventing cancer, comprisingadministering to a patient in need an effective amount of an antibody orfragment according to claim
 244. 253. The method of claim 252 furthercomprising co-administering a chemotherapeutic agent.
 254. The method ofclaim 252, wherein the cancer is selected from a group consisting ofhematological malignancies such as acute lymphocytic leukemia, chroniclymphocytic leukemia, acute myelogenous leukemia, chronic myelogenousleukemia, multiple myeloma, Hodgkin's lymphoma, Non-Hodgkin's lymphoma,and soft or solid tumors such as cancer of breast, prostate, lung,ovary, colon, spleen, kidney, bladder, head and neck, uterus, testicles,stomach, cervix, liver, bone, skin, pancreas, brain and wherein thecancer is non-metastatic, invasive or metastatic.
 255. The method ofclaim 252, wherein said cancer is selected from the group consisting oflung cancer, colon cancer, and wherein the lung cancer, or the coloncancer is non-metastatic, invasive or metastatic.
 256. The method ofclaim 255, wherein the lung cancer is a small cell lung cancer.
 257. Amethod for treating or preventing an immune disorder, comprisingadministering to a patient a therapeutically effective amount of anantibody according to claim
 244. 258. The method of claim 257, whereinthe treatment is combined with a moiety useful for treating said immunerelated condition.
 259. The method of claim 258, wherein the moiety is acytokine antibody, cytokine receptor antibody, drug, or anotherimmunomodulatory agent.
 260. The method of claim 259, wherein the immunerelated condition is selected from the group consisting of an autoimmunedisease, transplant rejection and graft versus host disease.
 261. Themethod of claim 260 wherein the autoimmune disease is selected from thegroup consisting of multiple sclerosis; psoriasis; rheumatoid arthritis;systemic lupus erythematosus; Crohn's disease, ulcerative colitis;immune disorders associated with graft transplantation rejection, benignlymphocytic angiitis, lupus erythematosus, Hashimoto's thyroiditis,primary myxedema, Graves' disease, pernicious anemia, autoimmuneatrophic gastritis, Addison's disease, insulin dependent diabetesmellitis, Good pasture's syndrome, myasthenia gravis, pemphigus,sympathetic ophthalmia, autoimmune uveitis, autoimmune hemolytic anemia,idiopathic thrombocytopenia, primary biliary cirrhosis, chronic actionhepatitis, ulceratis colitis, Sjogren's syndrome, rheumatic disease,polymyositis, scleroderma, mixed connective tissue disease, inflammatoryrheumatism, degenerative rheumatism, extra-articular rheumatism,collagen diseases, chronic polyarthritis, psoriasis arthropathica,ankylosing spondylitis, juvenile rheumatoid arthritis, periarthritishumeroscapularis, panarteriitis nodosa, progressive systemicscleroderma, arthritis uratica, dermatomyositis, muscular rheumatism,myositis, myogelosis and chondrocalcinosis.
 262. An assay for detectingthe presence of polypeptide as set forth in any one of SEQ ID NO:147,SEQ ID NO:148, SEQ ID NO:299, SEQ ID NO:48, or SEQ ID NO:50 in abiological sample comprising contacting the sample with an antibodyaccording to claim 244, and detecting the binding of SEQ ID NO:147, SEQID NO:148, SEQ ID NO:299, SEQ ID NO:48, or SEQ ID NO:50 in the sample.263. An isolated polynucleotide, comprising an amplicon having a nucleicacid sequence selected from the group consisting of 235, 238, 241, 244,or polynucleotides homologous thereto.
 264. An isolated polypeptideselected from H19011_(—)1_P8 (SEQ ID NO:48), H19011_(—)1_P9 (SEQ IDNO:50), or variant thereof that possesses at least 95% sequence identitytherewith.
 265. A nucleic acid sequence encoding a polypeptide accordingto claim
 264. 266. The nucleic acid sequence according to claim 265which is selected from the group consisting of H19011_(—)1_T8 (SEQ IDNO:45), H19011_(—)1_T9 (SEQ ID NO:46), or variant thereof that possessesat least 95% sequence identity therewith.